Development of a Method and Validation for the Quantitation of FruArg in Mice Plasma and Brain Tissue Using UPLC-MS/MS.
ABSTRACT: Aged garlic extract (AGE) is a popular nutritional supplement and is believed to promote health benefits by exhibiting antioxidant and anti-inflammatory activities and hypolipidemic and antiplatelet effects. We have previously identified N-?-(1-deoxy-d-fructos-1-yl)-l-arginine (FruArg) as a major contributor to the bioactivity of AGE in BV-2 microglial cells whereby it exerted a significant ability to attenuate lipopolysaccharide-induced neuroinflammatory responses and to regulate the Nrf2-mediated antioxidant response. Here, we report on a sensitive ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) protocol that was validated for the quantitation of FruArg in mouse plasma and brain tissue samples. Solid-phase extraction was used to separate FruArg from proteins and phospholipids present in the biological fluids. Results indicated that FruArg was readily absorbed into the blood circulation of mice after intraperitoneal injections. FruArg was reliably detected in the subregions of the brain tissue postinjection, indicating that it penetrates the blood-brain barrier in subnanomolar concentrations that are sufficient for its biological activity.
Project description:Objective:This study aimed to investigate the treatment effects of acupoint application of sinomenine in rheumatoid arthritis (RA). RA models were constructed using male New Zealand rabbits. Methods:The rabbits were randomly divided into a blank control group and four experimental groups as follows: ST 36 group (acupoint application of sinomenine at Zusanli); GB 34 group (acupoint application of sinomenine at Yanglingquan); knee-joint group (application directly at the site of the knee joint); and oral administration group (sinomenine administered by gavage). In all rabbits, microdialysis was applied at the knee joint to obtain samples. Pharmacokinetic (PK) and pharmacodynamic (PD) parameters were measured by ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), and the PK/PD models were established according to the parameters derived. Results:Sinomenine concentration was in the range of 0.832-208?ng/mL, and the peak area showed a good linear relationship with the regression equation of y?=?539.64x?+?953.81; r?=?0.9998. Moreover, good specificity and precision were obtained for the LC-MS/MS method of sinomenine evaluation in the microdialysate samples. The PK analysis showed that the sinomenine effect time was longer in the ST 36 group (area under the time-concentration curve (AUC): 12683.81?h·ng/ml and T max: 6.21?h) than in the other groups. Arginine and citrulline were selected as the indices for PD, and in the analysis of parameters for PK/PD, the highest value of E max and the lowest value of k e0 were obtained in the ST 36 group. Conclusion:Acupoint application of sinomenine at ST 36 has potential for use in patients with RA by enabling enhanced and prolonged treatment effects.
Project description:As a traditional Chinese medicine, Drynariae rhizoma (Kunze ex Mett.) J. Sm. has been used to treat osteoporosis and bone resorption for 2500 years. Based on the previous study and literature references, flavonoids were proved to be the most abundant and main active compounds of Drynariae rhizoma for osteoporosis treatment. In order to make good and rational use of Drynariae rhizoma in future, a rapid, sensitive, and selective ultraperformance liquid chromatography-mass spectrometry (UPLC-MS/MS) method was developed to investigate the pharmacokinetics of eight main flavonoids in rat plasma after oral administration of the Drynariae rhizoma extract, including neoeriocitrin, luteolin-7-O-?-D-glucoside, astragalin, naringin, eriodictyol, luteolin, naringenin, and kaempferol. Plasma samples' pretreatment involved a solid-phase extraction column. The separation was performed on an ACQUITY UPLCTM BEH C18 column with a gradient mobile-phase system of acetonitrile and 1% acetic acid in water. The detection was performed using a triple quadrupole tandem mass spectrometer equipped with an electrospray ionization interface (ESI) by multiple reaction monitoring (MRM) in the positive ion mode. All calibration curves exhibited good linearity (r 2 > 0.9990) over the measured ranges. The intraday and interday precisions (RSD) were within 13.87%, and the accuracy (RE) ranged from -14.57% to -0.25% at three quality control levels. Extraction recovery, matrix effect, and stability were satisfactory. The pharmacokinetic characteristics of the eight flavonoids of interest were clearly elucidated.
Project description:A selective, sensitive, and rapid method by using ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) for the determination of histamine in fish and fish sauce was developed. The optimal conditions of liquid chromatographic separation and mass spectroscopy of histamine have also been investigated. The linear ranges of the method were 20.0?÷?1000?ng/mL, and the corresponding correlation coefficient was 0.9993. Mean recoveries of the analyte at three spike levels (low, medium, and high) were within the range of 98.5%?÷?102.5% (n?=?7). The limit of detection (LOD) and limit of quantification (LOQ) values were 3.83 and 11.50?ng/mL for the fish sauce sample and 4.71 and 14.12?ng/mL for the fish sample, respectively. The influence of the matrix effect on the accuracy, repeatability, and recovery of the method was negligible. The recommended method was applied to determine the content of this substance in 21 fish sauce samples and 4 kinds of fish samples, which were collected from Ho Chi Minh City, Vietnam, in 2019.
Project description:Large quantities of thinned young pears, a natural source of bioactive compounds, are abandoned as agricultural by-products in many orchards. Hence, ten thinned young pear varieties were systematically investigated in terms of their chemical composition and antioxidant potential. Through ultra-performance liquid chromatography coupled with electrospray ionization triple quadrupole mass spectrometry (UPLC-Q TRAP-MS/MS), 102 polyphenols and 16 triterpenoids were identified and individually quantified within a short time using multiple reaction monitoring (MRM). Subsequently, the antioxidant capacities of these pears were determined with DPPH assays, and the correlation between total antioxidant activity and each component was analyzed. The results indicated that the bioactive compound content and antioxidant capacity in thinned pears were considerably high. Regarding chemical composition, chlorogenic acid, quinic acid and arbutin were the primary polyphenols and ursolic acid was the predominant triterpenoid, whereas 27 polyphenolic compounds, especially chlorogenic acid and most of the flavan-3-ols, were the main antioxidants in young pears. These findings should provide a scientific basis for the further use of pear fruit by-products.
Project description:Oxidative stress plays a role in the onset and progression of a number of diseases, such as Alzheimer's disease, diabetes and cancer, as well as ageing. Oxidative stress is caused by an increased production of reactive oxygen species and reduced antioxidant activity, resulting in the oxidation of glutathione. The ratio of reduced to oxidised glutathione is often used as a marker of the redox state in the cell. Whereas a variety of methods have been developed to measure glutathione in blood samples, methods to measure glutathione in cultured cells are scarce. Here we present a protocol to measure glutathione levels in cultured human and yeast cells using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC⁻MS/MS).
Project description:Annonaceous acetogenins (AAGs) are environmental neurotoxins from the fruit pulp of several Annonaceae species, whose consumption was linked to the occurrence of sporadic atypical Parkinsonism with dementia. The quantification of the prototypical AAG annonacin in Rat brain homogenates was performed by UPLC-MS/MS in selected reaction monitoring (SRM) mode, using a triple quadrupole mass analyzer. A natural analog of annonacin was used as an internal standard. Analyzed data set of the analytical validation of this method is presented, including stability of the samples, extraction recovery and matrix effect, supporting the results described in the article "Quantification of the environmental neurotoxin annonacin in Rat brain by UPLC-MS/MS" N. Bonneau, I. Schmitz-Afonso, D. Touboul, A. Brunelle, P. Champy (2016) .
Project description:Nuciferine is an aporphine alkaloid monomer that is extracted from the leaves of the lotus species <i>Nymphaea caerulea</i> and <i>Nelumbo nucifera</i> Gaertn. Nuciferine was reported to treat cerebrovascular diseases. However, the potential mechanism of the neuroprotective effects of nuciferine at the metabolomics level is still not unclear. The present research used neurological score, infarct volume, cerebral water content, and ultraperformance liquid chromatography to quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS)-based serum metabolomics to elucidate the anti-ischemic stroke effect and mechanisms of nuciferine. The results showed that nuciferine significantly improved neurological deficit scores and ameliorated cerebral edema and infarction. Multivariate data analysis methods were used to examine the differences in serum endogenous metabolism between groups, and the biomarkers of nuciferine on ischemic stroke were identified. Approximately 19 metabolites and 7 metabolic pathways associated with nuciferine on treatment of stroke were found, which indicated that nuciferine exerted a positive therapeutic action on cerebral ischemic by regulating metabolism. These results provided some data support for the study of anti-stroke effect of nuciferine from the perspective of metabolomics.
Project description:Flavonoids are a kind of essential substance for the human body because of their antioxidant properties and extremely high medicinal value. Citrus reticulata "Dahongpao" (DHP) is a special citrus variety that is rich in flavonoids, however little is known about its systematic flavonoids profile. In the present study, the presence of flavonoids in five important citrus varieties, including DHP, Citrus grandis Tomentosa (HZY), Citrus ichangensis Swingle (YCC), Citrus sinensis (L.) Osbeck (TC), and Citrus reticulata 'Buzhihuo' (BZH), was determined using a UPLC-ESI-MS/MS-based, widely targeted metabolome. Results showed that a total of 254 flavonoid metabolites (including 147 flavone, 39 flavonol, 21 flavanone, 24 anthocyanins, 8 isoflavone, and 15 polyphenol) were identified. The total flavonoid content of peels from DHP was the highest. DHP could be clearly separated from other samples through clustering analysis and principal component analysis (PCA). Further, 169 different flavonoid metabolites were observed between DHP peels and the other four citrus peels, and 26 down-regulated differential metabolites displayed important biological activities in DHP. At the same time, a unique flavonoid component, tricin 4'-O-syringyl alcohol, was only found in DHP, which could be used as a marker to distinguish between other varieties. This work might facilitate a better understanding of flavonoid metabolites between DHP peels and the other four citrus peels and provide a reference for its sufficient utilization in the future.
Project description:Background and Aim:Platycodon grandiflorum (PG) has been widely used for treating chronic bronchitis (CB). However, the material basis and underlying mechanism of action of PG against CB have not yet been elucidated. Methods:To analyze the ingredients in PG, ultraperformance liquid chromatography-quadrupole-time-of-flight tandem mass (UPLC-Q-TOF-MS/MS) technology was performed. Subsequently, using data mining and network pharmacology methodology, combined with Discovery Studio 2016 (DS), Cytoscape v3.7.1, and other software, active ingredients, drug-disease targets, and key pathways of PG in the treatment of CB were evaluated. Finally, the reliability of the core targets was evaluated using molecular docking technology and in vitro studies. Results:A total of 36 compounds were identified in PG. According to the basic properties of the compounds, 10 major active ingredients, including platycodin D, were obtained. Based on the data mining approach, the Traditional Chinese Medicine Systems Pharmacology Database, and the Analysis Platform (TCMSP), GeneCards, and other databases were used to obtain targets related to the active ingredients of PG and CB. Network analysis was performed on 144 overlapping gene symbols, and twenty core targets, including interleukin-6 (IL-6) and tumor necrosis factor (TNF), which indicated that the potential signaling pathway that was most relevant to the treatment of CB was the IL-17 signaling pathway. Conclusion:In this study, ingredient analysis, network pharmacology analysis, and experiment verification were combined, and revealed that PG can be used to treat CB by reducing inflammation. Our findings provide novel insight into the mechanism of action of Chinese medicine. Furthermore, our data are of value for the research and development of novel drugs and the application thereof.
Project description:Garlic and formulations containing allicin are used widely as fungicides in modern agriculture. However, limited reports are available on the allelopathic mechanism of green garlic volatile organic compounds (VOCs) and its component allelochemicals. The aim of this study was to investigate VOCs of green garlic and their effect on scavenging of reactive oxygen species (ROS) in cucumber. In this study, green garlic VOCs were collected by HS-SPME, then analyzed by GS-MS. Their biological activity were verified by bioassays. The results showed that diallyl disulfide (DADS) is the main allelochemical of green garlic VOCs and the DADS content released from green garlic is approximately 0.08 mg/g. On this basis, the allelopathic effects of green garlic VOCs in vivo and 1 mmol/L DADS on scavenging of ROS in cucumber seedlings were further studied. Green garlic VOCs and DADS both reduce superoxide anion and increase the accumulation of hydrogen peroxide of cucumber seedlings. They can also regulate active antioxidant enzymes (SOD, CAT, POD), antioxidant substances (MDA, GSH and ASA) and genes (CscAPX, CsGPX, CsMDAR, CsSOD, CsCAT, CsPOD) responding to oxidative stress in cucumber seedlings.