Mass Spectrometry Method Research Articles

In-silico based Designing of benzo[d]thiazol-2-amine Derivatives as Anal-gesic and Anti-inflammatory Agents.

Benzo[d]thiazoles represent a significant class of heterocyclic com-pounds renowned for their diverse pharmacological activities, including analgesic and anti-inflammatory properties. This molecular scaffold holds substantial interest among medicinal chemists owing to its structural versatility and therapeutic potential. Incorporating the benzo[d]thiazole moiety into drug molecules has been extensively investigated as a strategy to craft novel therapeutics with heightened efficacy and minimized adverse effects. The aim of the present research work was to design, synthesize and characterize the new benzo[d]thiazol-2-amine derivatives as potent analgesic and anti-inflammatory agents. The synthesis of the presented benzo[d]thiazol-2-amine derivatives was performed by condensing-(4-chlorobenzylidene) benzo[d]thiazol-2-amine with a number of substituted phenols in the presence of potassium iodide and anhydrous potassium carbonate in dry acetone. IR spectroscopy, 1HNMR spectroscopy, 13CNMR spectroscopy and Mass spectroscopy methods were used to characterize the structural properties of all 13 newly syn-thesized derivatives. The molecular properties of these newly synthesized derivatives were estimated to study the attributes of drug-like candidates. Benzo[d]thiazol-2-amine derivatives were molecularly docked with selective enzymes COX-1 and COX-2. Analgesic and anti-inflammatory activities of synthesized compounds were evaluated by us-ing albino rats. Findings of the research suggested that compounds G3, G4, G6, G8 and G11 possess higher binding affinity than diclofenac sodium, when docking was performed with enzyme COX-1. Compounds G1, G3, G6, G8andG10 showed lower binding affinity than Indometha-cin when docking was performed with enzyme COX-2.In vitro evaluation of the COX-1 and COX-2 enzyme inhibitory activities was performed for synthesized compounds. Compounds G10 and G11 exhibited significant COX-1 and COX-2 enzyme in-hibitory action with an IC50 value of 5.0 and 10 μM, respectively. Using the hot plate method and the carrageenan-induced rat paw edema model, the synthesized compounds were screened for their biological activities, including analgesic and anti-inflammatory activities. Highest analgesic action was exhibited by derivative G11 and the compound G10 showed the highest anti-inflammatory response. Inhibition of COX may be considered as a mechanism of action of these compounds. It was concluded that synthesized derivatives G10 and G11 exhibited significant analgesic and anti-inflammatory effect; therefore, the said compounds may be subjected to further clinical investigation for establishing these as future compounds for the treatment of pain and inflammation.

Continuous fiber printing of a modular heater for ion mobility spectrometry

Ion mobility spectrometry (IMS) is a powerful analytical method for separating and detecting gaseous substances like volatile organic compounds (VOCs). It is highly versatile, used both as a standalone technique and in combination with chromatographic methods for pre-separation or mass spectrometry for complex matrix analysis. This study explores the use of an advanced 3D printing technology to print a heater for an IMS-device.A continuous fiber printer, commonly used to enhance the mechanical properties of printed objects, was used to create a customized heating element by embedding a copper wire within the polymer matrix during the printing process. Cyclic olefin copolymer (COC) was chosen as the matrix material due to its analytical properties, such as low off-gassing and higher thermal stability, compared to the more commonly used polylactic acid (PLA).The 3D printed heater was incorporated into a drift tube ion mobility spectrometer, and its performance was evaluated by analyzing the separation of various ketones. The results show that the 3D printed heater effectively controlled the temperature within the drift tube, improving the resolving power of overlapping peaks.This study shows the potential of 3D printing technology to improve fabrication and functionality of IMS devices. The flexibility in design and material selection afforded by 3D printing not only enhances the analytical performance of IMS but also allows for customization and rapid prototyping of other analytical instruments or laboratory devices.

Preparation and evaluation of Puerarin-loaded PLGA nanoparticles for improving oral bioavailability in SD rats

BackgroundPuerarin (Pue) is an isoflavone compound with significant therapeutic effect on cardiovascular diseases, but its poor water solubility and low oral bioavailability limit clinical application. MethodsIn this study, Pue was prepared into PLGA nanoparticles (Pue-PLGA NPs) by emulsion solvent volatilization method. The morphology, particle size, Zeta potential, X-ray diffraction (XRD), and fourier transform infrared (FTIR) of the NPs were characterized. Additionally, their stability and in vitro release were evaluated. SD rats were orally administered wtih Pue and Pue-PLGA NPs, and a high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established to determine the concentration of blood samples and to investigate the pharmacokinetic behaviour of Pue and Pue-PLGA NPs in rats. ResultsThe NPs were observed by transmission electron microscopy (TEM) as regular spheroids and uniformly dispersed. The average particle size of the NPs was (167.1±5.26）nm, the Zeta potential was (-29.88±2.46)mV, the encapsulation rate was (83.12 %±4.73 %) and the drug loading capacity was (7.75 %±1.81 %). The results of in vitro release showed that the drug was released slowly and continuously from the NPs, reaching the release platform in 24 h, and the cumulative release amount was (88.55±2.86) %. The pharmacokinetic results showed that the AUC0–24, AUC0-∞, Cmax, Tmax, t1/2, MRT0–24 and MRT0-∞ of Pue-PLGA NPs were 2.196, 1.978, 1.327, 1.5, 1.385, 3.915 and 3.140 times of Pue, respectively. The relative bioavailability was (197.82±25.28) %. ConclusionThese results indicated that the prepared nanoparticles had small particle size, high encapsulation rate, drug loading capacity and good slow-release effect, and could significantly improve the oral bioavailability of Pue in rats.

Integration of ion mobility-quadrupole time-of-flight mass spectrometry and triple quadrupole-ion trap mass spectrometry coupled with ultra-high performance liquid chromatography for characterizing and quantitatively assay the saponins of Panax quinquefolius flower

Panax quinquefolius (American ginseng) is globally popular serving as herbal medicine or tonifying agent. Compared with the root, studies on the flower of Panax quinquefolius (PQF) in respect of the multicomponent characterization and quality evaluation is scarce. This work was designed, by feat of two liquid chromatography–mass spectrometry platforms, to characterize and quantitatively assay the saponins of PQF. Ultra-high performance liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (UHPLC/IM-QTOF-MS) combined with data-independent high-definition MSE (HDMSE) was developed to acquire the structure information of PQF saponins. Consequently, efficient and more accurate identification (higher identification accuracy and much lower false positives rate) of the involved saponins were accomplished by the intelligent peak annotation workflows of the UNIFI bioinformatics platform incorporated with a “Multi-dimensional Information Library of Ginsenosides” (GinMIL). Totally 128 ginsenosides were identified or tentatively characterized, and 45 thereof were identified by comparing with the reference standards. Additionally, an efficient UHPLC/QTrap-MS (triple quadrupole-ion trap mass spectrometry) method enabling quantitative assay of 21 ginsenosides in PQF was established and validated by scheduled multiple reaction monitoring (sMRM). It gave low limit of detection (0.011–0.65 pg) and limit of quantification (0.053–4.1 pg), good linearity (R2 > 0.99), intra-/inter-day precision (RSD < 13.6 %), stability (RSD < 3.5 %), repeatability (RSD < 7.5 %), and recovery (RSD < 13.5 %). It application to quantify 21 ginsenosides in 14 batches of PQF samples revealed differences in their saponin content. Conclusively, systematic multi-component characterization and multi-indicator quantitation of PQF were achieved, thus providing the methodological reference for the quality investigation of functional herbs.

Locations of Functional Groups in the Aliphatic Chains of Lipids and the Arrangement of Aliphatic Chains within Acylglycerolipids by the Methods of Mass Spectrometry

Locations of Functional Groups in the Aliphatic Chains of Lipids and the Arrangement of Aliphatic Chains within Acylglycerolipids by the Methods of Mass Spectrometry

Microbiota of complete removable dentures

Purpose: to study the quantitative and qualitative composition of microflora on the surface of complete removable dentures after 1 and 4 years of use. Materials and methods: 40 patients were diagnosed with complete edentulism (K08.1) using complete removable acrylic dentures for no more than 4 years were examined. The composition of the microflora was determined by mass spectrometry method. Statistical processing of the results of the study was carried out using the computer package Statistica13. To assess the differences between the groups, parametric t-criteria with Bonferroni correction were used for multiple comparisons, the critical level of statistical significance of p was 0.05. Results: it was found that in 4 years the number of yeast and fungi increased by 91%, n=20 (100%); the number of actinobacteria increased by 61%, n=20 (100%); the number of gram-negative bacteria increased by 500% n=20 (100%); the number of anaerobic microorganisms increased by 50% n=20 (100%); the number of cocci and bacilli increased by 190% n=20 (100%). Conclusion: after conducting the research, we found that after 4 years of using removable dentures n=20 (100%), the number of cocci and bacilli increased from 56±5 (105 cells/gram) (p=0.03) to 107±8 (105 cells/gram) (p=0.04). The number of anaerobes increased n=20 (100%) from 68±6 (105 cells/gram) (p=0.0002) to 102±9 (105 cells/gram) (p=0.0002). The number of actinobacteria n=20 (100%) increased from 30±3 (105 cells/gram) to 143±12 (105 cells/gram) (p=0.003). The number of gram-negative bacteria n=20 (100%) increased from 4±1 (105 cells/gram) (p=0.0005) to 24±2 (105 cells/gram) (p=0.0006). The number of fungi and yeast n=20 (100%) increased from 977±90 (105 cells/gram) (p=0.0003) to 1587±136 (105 cells/gram) (p=0.003).

Detection of "smoke powder" etomidate and its metabolite etomidate acid in blood and urine by UHPLC-MS-MS: application in authentic cases.

Recently, etomidate has been widely used as an alternative in illicit drug market. It is usually added to regular cigarette tobacco (commonly known as "cigarette powder") or mixed in e-cigarette oil sold through the Internet, retail stores, or entertainment outlets and other channels. An ultra-high performance liquid chromatography tandem mass spectrometry method was developed to quantify etomidate and etomidate acid in human blood and urine. The limit of detection (LOD) of etomidate and etomidate acid in blood is 0.5 and 2 ng/mL, respectively, and the lower limit of quantification (LLOQ) is 1 and 5 ng/mL, respectively. The LOD of etomidate and etomidate acid in urine is 1 and 2 ng/mL, respectively, and the LLOQ is 2 and 5 ng/mL, respectively. The precision, accuracy, recoveries, and matrix effects of etomidate and etomidate acid determinations in blood and urine met the requirements for methodological validation. The method was successfully applied to the identification and quantification of etomidate and etomidate acid in blood and urine of 62 forensic cases. The concentration of etomidate ranged from 1.52 to 8.41 ng/mL (positive cases, n = 5) and the concentration of etomidate acid ranged from 2.76 to 112 ng/mL (positive cases, n = 5) in blood. The concentrations of etomidate and etomidate acid in urine samples were 2.64-79,300 ng/mL (positive cases, n = 59) and 6.11-518,000 ng/mL (positive cases, n = 60), respectively. Therefore, the concentration of etomidate in blood and urine is mostly higher than that of etomidate.

Development and Clinical Validation of Liquid Chromatography-Tandem Mass Spectrometry for Measuring Ruxolitinib in Steroid-Refractory Graft-Versus-Host Disease: A First Step Towards Optimized Treatment.

This non-interventional, prospective, single-center study aimed to develop a technique to measure ruxolitinib (RUX) concentrations and provide preliminary data on the distribution of plasma drug levels in patients with steroid refractory (SR) GvHD. Between April 2023 and May 2024, we analyzed 48 blood samples from 29 patients with SR-GvHD. Median individual plasma concentrations varied across different RUX doses and largely overlapped: 39.2 ng/mL at 10 mg b.i.d (range: 0-73), 13.1 ng/mL at 10-5 mg (range, 6.1-35.6), and 31.6 ng/mL at 5 mg b.i.d (range: 0.7-99.9). Samples taken under non-optimal temperature conditions showed a lower median concentration of 0.77 ng/mL (range: 0-7.4 ng/mL). The four patients who did not respond at days +28 and +180 after RUX initiation (3 with lower gastrointestinal aGvHD, and 1 with ocular, hepatic, and pulmonary cGvHD) showed a median concentration of only (7.4 ng/mL (range, 0-29) ng/mL) with full dosing. The introduction and validation of a liquid chromatography-tandem mass spectrometry method for quantifying plasma RUX concentrations was feasible in our center. Administering predetermined and fixed doses of RUX in patients with SR-GvHD showed highly variable and overlapping plasma drug concentrations. This underscores the potential importance of RUX- pharmacokinetics (PK) monitoring.

Bubble-Assisted Sample Preparation Techniques for Mass Spectrometry.

This review delves into the efficacy of utilizing bubbles to extract analytes into the gas phase, offering a faster and greener alternative to traditional sample preparation methods for mass spectrometry. Generating numerous bubbles in liquids rapidly transfers volatile and surface-active species to the gas phase. Recently, effervescence has found application in chemical laboratories for swiftly extracting volatile organic compounds, facilitating instantaneous analysis. In the so-called fizzy extraction, liquid matrices are pressurized with gas and then subjected to sudden decompression to induce effervescence. Alternatively, specifically designed effervescent tablets are introduced into the liquid samples. In situ bubble generation has also enhanced dispersion of extractant in microextraction techniques. Furthermore, droplets from bursting bubbles are collected to analyze non-volatile species. Various methods exist to induce bubbling for sample preparation. The polydispersity of generated bubbles and the limited control of bubble size pose critical challenges in the stability of the bubble-liquid interface and the ability to quantify analytes using bubble-based sample preparation techniques. This review covers different bubble-assisted sample preparation methods and gives practical guidance on their implementation in mass spectrometry workflows. Traditional, offline, and online approaches for sample preparation relying on bubbles are discussed. Unconventional bubbling techniques for sample preparation are also covered.

Rapid Untargeted Puff‐by‐Puff Analysis of (Electronic) Cigarette Emissions by Concentric Dielectric Barrier Discharge Ionisation Mass Spectrometry

AbstractDespite the soaring popularity of e‐cigarettes among teenagers and young adults, our understanding of the full extent of their health hazards have remained limited. This is due to the vast complexities of e‐cigarette aerosols and the difficulty in their full characterisation. Conventional mass spectrometry methods of e‐cigarette analysis, though pioneering in driving political and medical discourse, have been limited in their capabilities to uncover all compounds in its emissions due to prominent limitations in experimental setup. To overcome this major hurdle, we have developed a setup for puff‐by‐puff analysis of electronic and conventional cigarette emissions by concentric dielectric barrier discharge ionisation mass spectrometry. In this pilot study, the simple setup of in‐line dilution and high‐resolution mass spectrometry analysis allowed us to directly uncover 225 compounds in e‐cigarette puffs across a wide spectrum of chemical classes in two sequential 5‐minute runs. These include acids, carbonyls, aromatic cyclics, heterocyclics, unsaturated and saturated hydrocarbons, alcohols, esters, alkaloids, sulfur‐containing compounds, oxides, and nitriles. As a result, our setup provided a significant improvement in rapid compound identification, and demonstrated a much broader chemical landscape in e‐cigarette emissions than previously reported.

Impact of Citrate on Mitigating Iron Mediated Polysorbate 80 Degradation in Biotherapeutic Formulation Placebos.

Polysorbate 80 (PS80), a widely used polymeric surfactant in biotherapeutic formulation, possesses a unique structural composition that effectively prevents protein aggregation in highly concentrated protein drug formulations. However, PS80 is susceptible to hydrolysis, due to the presence of fatty acid esters that can be enzymatically hydrolyzed, The unsaturated bonds in the fatty acids are prone to oxidative degradation when exposed to air, especially in the presence of transition metals such as iron and copper, which may be introduced during production and purification processes or from contamination in raw materials used in drug formulation. The degradation of PS80, particularly through metal-mediated oxidative degradation, poses a significant challenge for the industry. Among the identified trace metals, iron plays a crucial role as the redox reaction between ferrous ion (Fe(II)) and ferric ion (Fe(III)) generates radicals that initiate the degradation process. In order to investigate the impact of iron on PS80 degradation and understand the mechanism of iron-catalyzed oxidation, we utilized charge-reduction mass spectrometry and two-dimensional ion density mapping technologies to characterize the degradation of PS80. This method has proven to be a convenient and effective tool for the quick and detailed profiling of PS80, allowing for visual monitoring and examination of the changes that reflect the difficult-to-identify and easy-to-miss oxidized species of PS80. Additionally, a high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry method was developed for the separation and measurement of Fe(II) and Fe(III). Through this investigation, we determined that the involvement of Fe(II)/Fe(III) in PS80 degradation is a temperature dependent process. Furthermore, we found citrate not only promotes the conversion of Fe(II) to Fe(III), but it also chelates Fe(III) and prevents its reduction to Fe(II), thus inhibiting the initiation of the PS80 degradation. Therefore, the addition of citrate can be a crucial ingredient for controlling the degradation of PS80 in biologic drug substances and products. Overall, this investigation has provided valuable insights to enhance product stability, optimize processes, and ensure the quality of formulations containing PS80.

Pharmacokinetics and Bioequivalence of Two Formulations of Montelukast Sodium Tablets in Healthy Chinese Volunteers Under Fasting and Fed Conditions.

Montelukast sodium is a leukotriene type 1 receptor antagonist that can be used for the prophylaxis and treatment of asthma. However, the pharmacokinetics of montelukast sodium tablets (10mg) remain unclear in healthy Chinese subjects. Here, a single-dose randomized, open-label, 2-sequence, and 2-period crossover (7-day washout period between treatments) study was performed to compare the pharmacokinetics and bioequivalence between the test products and the reference at a single dose of 10mg among healthy Chinese subjects under fasting and fed conditions. Blood samples were collected at specified time points to analyze the plasma concentrations of montelukast by a validated liquid chromatography-tandem mass spectrometry method. The results showed that the 90% confidence interval values of the geometric mean ratio of test/reference for the maximum plasma drug concentration, area under the concentration-time curve from time 0 to the end, and area under the concentration-time curve from time 0 to infinity were within the range of 80%-125%. Moreover, both the test and reference formulations were safe and well tolerated, with no occurrence of severe adverse events. These results demonstrate that both the test montelukast sodium tablets and the reference showed similar bioequivalence, safety, and tolerability among healthy Chinese subjects under fasting and fed conditions.

Development and validation of a rapid HPLC-MS/MS method for simultaneous determination of cyclosporine A and tacrolimus in whole blood for routine therapeutic drug monitoring in organ transplantation.

Therapeutic drug monitoring is an integral part of organ transplantation. A rapid, simple, economical, and robust high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for simultaneously determining the immunosuppressants cyclosporine A and tacrolimus might increase detection efficiency. In this study, we developed and validated a rapid HPLC-MS/MS method. Whole blood samples of 100 μL were prepared by protein precipitation with acetonitrile and 0.5mol. L-1 ZnSO4. Chromatography was performed on a pre-column using a gradient elution with 20mmol. L-1 ammonium formate and 0.1% (v/v) formic acid in water (mobile phase A) and 0.1% (v/v) formic acid in methanol (mobile phase B) at a flow rate of 1.5mL.min-1. The analysis time was 2.2min. Electrospray ionization and multiple reaction monitoring were performed. The lower limit of quantification was set at 1ng. L-1 for tacrolimus and 50 ng. L-1 for cyclosporine A. The method showed adequate accuracy and precision with a sufficient linear range. The calibration curve range of tacrolimus and cyclosporine A was 1-30 and 50-1500 ng·mL-1, respectively. All correlation coefficients were >0.99. The developed HPLC-MS/MS is rapid and can be used for simultaneous monitoring of tacrolimus and cyclosporine A.

Simultaneous Quantification of Serum Symmetric Dimethylarginine, Asymmetric Dimethylarginine and Creatinine for Use in a Routine Clinical Laboratory.

Background Symmetric dimethylarginine (SDMA) and asymmetric dimethylarginine (ADMA) are naturally occurring amino acids classed as uremic toxins by the EUTOX working group. SDMA is principally excreted through the kidneys and is a well-known renal function marker, ADMA is a potent inhibitor of nitric oxide production. Here, we describe the development of a rapid and sensitive liquid chromatography tandem mass spectrometry method for simultaneous measurement of SDMA, ADMA and creatinine. Method Serum samples were prepared by protein precipitation and dilution with acetonitrile prior to injection onto a Waters TQS-Micro. Multiple reaction monitoring was used to detect SDMA, ADMA, creatinine, and their corresponding internal standard transitions after separation with a HILIC analytical column. Sample stability and intra-individual variation studies were also assessed following ethical approval. Results The retention time for creatinine was 0.43, SDMA 1.10 and ADMA 1.14 minutes. Mean recovery for creatinine was 103%, SDMA was 100% and ADMA was 103%, matrix effects were minimal (<6%). Lower limit of quantitation for creatinine and SDMA/ADMA was 17.5 µmol/L and 0.1 µmol/L respectively. Analytical imprecision showed a coefficient of variation <10% for all analytes across the working range of the assays. Intra-individual variation for creatinine was 4.7%, SDMA 7.5% and ADMA 7.6%. Discussion We have developed a rapid assay for LC-MS/MS measurement of SDMA, ADMA and creatinine in a routine clinical laboratory. It is simple, reproducible, and easy to perform. The stability of SDMA and ADMA pre- and post-centrifugation allows for their routine use without any special sample handling requirements.

A Micro-Flow Liquid Chromatography-Mass Spectrometry Method for the Quantification of Oxylipins in Volume-Limited Human Plasma.

Oxylipins are well-known lipid mediators in various inflammatory conditions. Their endogenous concentrations range from low picomolar to nanomolar, and there are growing demands to determine their concentrations in low-volume matrices for pathological studies, including blood, cerebrospinal fluids from animal disease models, infants, and microsampling devices. Most of the published quantification methods for comprehensive profiling of oxylipins still require more than 50µL plasma as a starting volume to detect these low levels. The aim of our study is to develop a sensitive and reliable method for the quantification of oxylipins in volume-limited human plasma samples. We established and validated a micro-liquid chromatography (LC)-mass spectrometry (MS)/MS method that requires only 5µL of human plasma for the determination of 66 oxylipins. The optimized micro-LC-MS/MS method utilized a flow rate of 4µL/min with a 0.3-mm inner diameter column. With an injection volume of 3µL, our method provides limits of detection in the range from 0.1 to 91.9pM, and limits of quantification range from 0.3 to 306.2pM. The sensitivity enhancement compared to conventional flow ranged from 1.4 to 180.7 times for 51 compounds depending on their physical-chemical properties. After validation, the method was applied to analyze 40 plasma samples from a healthy aging study to demonstrate robustness and sensitivity.

Provenance and Paleo-Environment of the Late Carboniferous Bauxite Formations in Southern Shanxi

The Carboniferous Benxi Formation in southern Shanxi of North China has significant bauxite resource potential; however, the source of its metallogenic material and its sedimentary environment remain unclear. The microscopy, X-ray diffraction, X-ray fluorescence spectroscopy, and inductively coupled plasma mass spectrometry methods were applied in this study to examine the mineralogical, petrographic, and geochemical characteristics. Geochemical proxies of La/Y, Sr/Ba, Al2O3/TiO2, Zr/Sc, Th/Sc, La/Sc, and Th/Co were analyzed to investigate the paleo-depositional environment and provenance of the aluminum-bearing strata. The findings indicate that diaspores are the primary ore minerals in bauxite, while kaolinite and rutile are the predominant gangue minerals. Both the bauxite and claystone/aluminous rocks exhibit high enrichment in Li, Bi, and U, with relative enrichment in In, Sb, Th, Nb, and Ta. Li is notably concentrated in the claystone/aluminous rocks, reaching up to 1994.00 μg/g, primarily occurring in cookeite and boehmite, while U is highly concentrated in the bauxite. The aluminum-bearing strata were primarily formed under alkaline-reducing conditions, with changes in acidity and alkalinity of the environment during the sedimentary diagenetic process. Marine transgressions significantly impacted the sedimentary environment of the aluminum-bearing strata, and the paleoclimate was characterized as hot and humid. The principal factors contributing to enrichment of aluminum in the sedimentary basin were the in situ weathering of aluminum-rich source rocks and the transport of clastic materials from high-aluminum source rocks. The source rocks were closely associated with intermediate-acidic magmatic rocks and potentially related to the weathering of Ordovician carbonates.

Bioactivity-Based Analysis and Chemical Characterization of Hypoglycemic Components from Helicteres angustifolia L.

Helicteres angustifolia L. (H. angustifolia), a well-known traditional Chinese medicine, has been demonstrated to have hypoglycemic activity. We found that the EtOAc extract of H. angustifolia (HAEF) showed stronger α-glucosidase inhibitory activity than that of positive control. Furthermore, the hypoglycemic activity of HAEF was evaluated in streptozotocin (STZ)-induced type 2 diabetes mellitus (T2DM) rats. The results demonstrated that HAEF reduced the drinking quantity, feeding quantity, and controlled weight loss in diabetic rats. Besides, the fasting blood glucose (FBG), viscera index, and the area under time-blood glucose curve (AUC) were significantly decreased, and the oral glucose tolerance was also improved after 5 weeks. Then, the high-performance liquid chromatography with quadrupole time of flight tandem mass spectrometry (HPLC-Q-TOF-MS/MS) method was performed for qualitative analysis of the chemical constituents in HAEF. Twenty-one compounds were identified from in HAEF. Four compounds were further isolated from HAEF and subjected to α-glucosidase inhibition experiments. At the end, molecular docking was empolyed simulate the interaction of three compounds with α-glucosidase. This is the first report on major hypoglycaemic components has been identified in the roots of H. angustifolia. These findings provide a material basis for the use of H. angustifolia in the treatment of diabetes.

Zic-HILIC MS/MS Method for NADomics Provides Novel Insights into Redox Homeostasis in Escherichia coli BL21 Under Microaerobic and Anaerobic Conditions

Background: Nicotinamide adenine dinucleotide (NAD+), its precursors, and its derivatives (collectively NADome) play a crucial role in cellular processes and maintain redox homeostasis. Understanding the dynamics of these metabolic pools and redox reactions can provide valuable insights into metabolic functions, especially cellular regulation and stress response mechanisms. The accurate quantification of these metabolites is challenging due to the interconversion between the redox forms. Methods: Our laboratory previously developed a zwitterionic hydrophilic interaction liquid chromatography (zic-HILIC)–tandem mass spectrometry method for the quantification of five essential pyridine nucleotides, including NAD+ derivatives and it’s reduced forms, with 13C isotope dilution and matrix-matched calibration. In this study, we have improved the performance of the chromatographic method and expanded its scope to twelve analytes for a comprehensive view of NAD+ biosynthesis and utilization. The analytical method was validated and applied to investigate Escherichia coli BL21 under varying oxygen supplies including aerobic, microaerobic, and anaerobic conditions. Conclusions: The intracellular absolute metabolite concentrations ranged over four orders of magnitude with NAD+ as the highest abundant, while its precursors were much less abundant. The composition of the NADome at oxygen-limited conditions aligned more with that in the anaerobic conditions rather than in the aerobic phase. Overall, the NADome was quite homeostatic and E. coli rapidly, but in a minor way, adapted the metabolic activity to the challenging shift in the growth conditions and achieved redox balance. Our findings demonstrate that the zic-HILIC-MS/MS method is sensitive, accurate, robust, and high-throughput, providing valuable insights into NAD+ metabolism and the potential significance of these metabolites in various biological contexts.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for rapid analysis of eight Gelsemium elegans Benth alkaloids in human plasma and urine

Gelsemium elegans Benth alkaloids are the main components of G. elegans and can cause acute toxicosis or even death. Although several studies have reported methods for detecting G. elegans alkaloids, a high-throughput and environmental-friendly strategy for detection of multiple G. elegans alkaloids has not been realized. In this work, a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry method was developed for rapid detection of G. elegans alkaloids in human plasma and urine for diagnosis of poisoning. Multiple matrices and crystal spotting methods were evaluated to obtain stable and high peak intensities without “sweet spot”. We verified the methodology and obtained excellent results. The matrix effects with different dilutions were compared and good recoveries and a low relative standard deviation were obtained with a 40-fold dilution. This method could shorten the analysis time and greatly reduce the consumption of chemical solvents. Furthermore, it could be applied to quantitative assessment of G. elegans alkaloid poisoning incidents.

A novel, sensitive, and fast ultra-high-performance liquid chromatography tandem mass spectrometry method for TNG908 determination in dog plasma and pharmacokinetic study.

TNG908 is a potent and selective protein arginase methyltransferase 5 (PRMT5) inhibitor that is currently going through phase I/II clinical development for the treatment of non-small cell lung cancer. To facilitate pharmacokinetic and toxicokinetic studies of TNG908, here, we reported an ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the detection of TNG908 in dogs. The dog plasma samples were precipitated by acetonitrile and analyzed using a Waters ACQUITY BEH C18 column combined with a Thermo triple quadrupole mass spectrometer. The mobile phase consisted of 0.1% formic acid solution and acetonitrile, at a flow rate of 0.3mL/min. TNG908 and internal standard were monitored by selective reaction monitoring (SRM) with m/z 410.2 > 150.1 and m/z 394.2 > 278.1, respectively. The method demonstrated excellent linearity over the concentration range of 1-1000 ng/mL, with a correlation coefficient greater than 0.995. Acetonitrile-mediated protein precipitation showed high extraction efficiency and a recovery above 80%. The validated assay was further applied to measure TNG908 in dog plasma after oral and intravenous administration and achieved success. The obtained pharmacokinetic parameters indicated low clearance of TNG908 (3.7 ± 0.8mL/min/kg) and moderate oral bioavailability (>36.4%).