[Analysis and comparison of metabolic processes of salvianolic acid A and salvianolic acid B in rats based on UHPLC-LTQ-Orbitrap MS technology].

The metabolites of salvianolic acid A and salvianolic acid B in rats were analyzed and compared by ultra-high-perfor-mance liquid chromatography with linear ion trap-orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS). After the rats were administrated by gavage, plasma at different time points and urine within 24 hours were collected to be treated by solid phase extraction(SPE), then they were gradient eluted by Acquity UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 µm) and 0.1% formic acid solution(A)-acetonitrile(B) mobile phase system, and finally all biological samples of rats were analyzed under negative ion scanning mode. By obtaining the accurate relative molecular mass and multi-level mass spectrometry information of metabolites, combined with the characteristic cleavage law of the reference standard and literature reports, a total of 30 metabolites, including salvianolic acid A and B, were identified. Among them, there were 24 metabolites derived from salvianolic acid A, with the main metabolic pathways including ester bond cleavage, dehydroxylation, decarboxylation, hydrogenation, methylation, hydroxylation, sulfonation, glucuronidation, and their multiple reactions. There were 15 metabolites of salvianolic acid B, and the main biotransformation pathways were five-membered ring cracking, ester bond cleavage, decarboxylation, dehydroxylation, hydrogenation, methylation, sulfonation, glucuronidation, and their compound reactions. In this study, the cross-metabolic profile of salvianolic acid A and B was elucidated completely, which would provide reference for further studies on the basis of pharmacodynamic substances and the exploration of pharmacological mechanism.

[Analysis of carnosic acid metabolites in rats by UHPLC-Q-Exactive MS].

A method of ultra-high performance liquid chromatography coupled with quadrupole/electrostatic field Obitrap high-resolution mass spectrometry(UHPLC-Q-Exactive MS) was established to comprehensively identify the metabolites of carnosic acid in rats. After oral gavage of carnosic acid CMC-Na suspension in rats, urine, plasma and feces samples were collected and pretreated by solid phase extraction(SPE). Acquity UPLC BEH C_(18 )column(2.1 mm×100 mm, 1.7 µm) was used with 0.1% formic acid solution(A)-acetonitrile(B) as the mobile phase for the gradient elution. Biological samples were analyzed by quadrupole/electrostatic field Obitrap high-resolution mass spectrometry in positive and negative ion mode. Based on the accurate molecular mass, fragment ion information, and related literature reports, a total of 28 compounds(including carnosic acid) were finally identified in rat samples. As a result, the main metabolic pathways of carnosic acid in rats are oxidation, hydroxylation, methylation, glucuronide conjugation, sulfate conjugation, S-cysteine conjugation, glutathione conjugation, demethylation, decarbonylation and their composite reactions. The study showed that the metabolism of carnosic acid in rats could be efficiently and comprehensively clarified by using UHPLC-Q-Exactive MS, providing a reference for clarifying the material basis and metabolic mechanism of carnosic acid.

Panax notoginseng Saponins Ameliorate Leukocyte Adherence and Cerebrovascular Endothelial Barrier Breakdown upon Ischemia-Reperfusion in Mice.

Panax notoginseng saponins (PNS) are known as clinical anti-stroke herbal medicines. The aim of this study is to describe the impact of PNS on ischemia-reperfusion-induced cerebral microvasculature barrier dysfunction which has not been investigated yet. Mice were subjected to transient middle cerebral artery occlusion and PNS were administrated to mice 3 days before and 2 days after surgery. Leukocyte adhesion, albumin leakage, tight junctions and other parameters in the cortex were measured. The PNS 45 mg/kg intervention alleviated leukocyte adhesion, inhibited endothelial barrier alterations evidenced by reduced albumin leakage and tight junction degradations, and ultimately ameliorated infarct volumes and neurological deficits subsequent to ischemia-reperfusion. Taken together, P. notoginseng saponins are able to attenuate leukocyte-mediated microvascular disturbance at the onset of ischemic stroke.

A Comprehensive Screening and Identification of Genistin Metabolites in Rats Based on Multiple Metabolite Templates Combined with UHPLC-HRMS Analysis.

Genistin, an isoflavone belonging to the phytoestrogen family, has been reported to possess various therapeutic effects. In the present study, the genistin metabolites in rats were investigated by UHPLC-LTQ-Orbitrap mass spectrometer in both positive and negative ion modes. Firstly, the data sets were obtained based on data-dependent acquisition method and then 10 metabolite templates were established based on the previous reports. Then diagnostic product ions (DPIs) and neutral loss fragments (NLFs) were proposed to efficiently screen and ascertain the major-to-trace genistin metabolites. Meanwhile, the calculated Clog P values were used to identify the positional isomers with different retention times. Consequently, a total of 64 metabolites, including prototype drug, were positively or putatively characterized. Among them, 40 metabolites were found according to the templates of genistin and genistein, which was the same as the previous research method. After using other metabolite templates, 24 metabolites were added. The results demonstrated that genistin mainly underwent methylation, hydrogenation, hydroxylation, glucosylation, glucuronidation, sulfonation, acetylation, ring-cleavage and their composite reactions in vivo biotransformation. In conclusion, the research not only revealed the genistein metabolites and metabolic pathways in vivo comprehensively, but also proposed a method based on multiple metabolite templates to screen and identify metabolites of other natural compounds.

Acid rain reduced soil carbon emissions and increased the temperature sensitivity of soil respiration: A comprehensive meta-analysis.

Soil respiration the second-largest carbon flux in terrestrial ecosystems, has been extensively studied across a wide range of biomes. Surprisingly, no consensus exist on how acid rain (AR) impacts the spatiotemporal pattern of soil respiration. Therefore, we conducted a meta-analysis using 318 soil respiration and 263 soil respiration temperature sensitivity (Q10) data points obtained from 48 studies to assess the impact of AR on soil respiration components and their Q10. The results showed that AR reduced soil total respiration (Rt) and soil autotrophic respiration (Ra) by 7.41 % and 20.75 %, respectively. As the H+ input increased, the response rates of Ra to AR (RR-Ra) and soil heterotrophic respiration (Rh) to AR (RR-Rh) decreased and increased, respectively. With increased AR duration, the RR-Ra increased, whereas the RR-Rh did not change. AR increased the Q10 of Rt (Rt-Q10) and Rh (Rh-Q10) by 1.92 % and 9.47 %, respectively, and decreased the Q10 of Ra (Ra-Q10) by 2.77 %. Increased mean annual temperature, mean annual precipitation, and initial soil organic carbon increased the response rate of Ra-Q10 to AR (RR-Ra-Q10) and decreased the response rate of Rh-Q10 to AR (RR-Rh-Q10). However, as the AR frequency and initial soil pH increased, both RR-Ra-Q10 and RR-Rh-Q10 also increased. In summary, AR decreased Rt but increased Q10, likely due to soil acidification (soil pH decreased by 7.84 %), reducing plant root biomass (decreased by 5.67 %) and soil microbial biomass (decreased by 5.67 %), changing microbial communities (increased fungi to bacteria ratio of 15.91 %), and regulated by climate, vegetation, soil and AR regimes. To the best of our knowledge, this is the first study to reveal the large-scale, varied response patterns of soil respiration components and their Q10 to AR. It highlights the importance of applying the reductionism theory in soil respiration research to enhance our understanding of soil carbon cycling processes with in the context of global climate change.

Comprehensive Screening and Identification of Phillyrin Metabolites in Rats Based on UHPLC-Q-Exactive Mass Spectrometry Combined with Multi-Channel Data Mining.

Phillyrin, a well-known bisepoxylignan, has been shown to have many critical pharmacological activities. In this study, a novel strategy for the extensive acquisition and use of data was established based on UHPLC-Q-Exactive mass spectrometry to analyze and identify the in vivo metabolites of phillyrin and to elucidate the in vivo metabolic pathways of phillyrin. Among them, the generation of data sets was mainly due to multichannel data mining methods, such as high extracted ion chromatogram (HEIC), diagnostic product ion (DPI), and neutral loss filtering (NLF). A total of 60 metabolites (including the prototype compound) were identified in positive and negative ion modes based on intuitive and useful data such as the standard's cleavage rule, accurate molecular mass, and chromatographic retention time. The results showed that a series of biological reactions of phillyrin in vivo mainly included methylation, hydroxylation, hydrogenation, sulfonation, glucuronidation, demethylation, and dehydrogenation and their composite reactions. In summary, this study not only comprehensively explained the in vivo metabolism of phillyrin, but also proposed an effective strategy to quickly analyze and identify the metabolites of natural pharmaceutical ingredients in nature.

Royal jelly increases peripheral circulation by inducing vasorelaxation through nitric oxide production under healthy conditions.

AIMS: Royal jelly (RJ) has a variety of reported biological activities, including vasorelaxation and blood pressure-lowering effects. Although functional foods are positively used for health, the effects of RJ on the cardiovascular system in healthy individuals have not been well studied. Therefore, we investigated the mechanisms underlying the vasorelaxation effects of RJ in healthy control rats to evaluate whether the peripheral circulation was increased. MAIN METHODS: We used fresh RJ to examine the vasorelaxation effects and related mechanisms in Wistar rats using organ bath techniques. Furthermore, we measured changes in tail blood circulation, systolic blood pressure (sBP), and heart rate (HR) after the oral administration of RJ to control rats and nitro-l-arginine methyl ester (l-NAME)-treated rats (0.5 mg/ml dissolved in distilled drinking water for 1 week). Concentrations of acetylcholine (ACh) in the RJ were measured using a commercial kit. KEY FINDINGS: RJ caused vasorelaxation of isolated rat aortas and superior mesenteric arteries, and this effect was inhibited by atropine (10-5 M, 15 min) or L-NAME (10-4 M, 20 min) and endothelium-denuded arterial ring preparations. Oral RJ increased tail blood flow and mass in control rats 1 h after treatment without affecting velocity, sBP, or HR. These effects were not observed in L-NAME-treated rats. RJ contained approximately 1000 µg/g of ACh. SIGNIFICANCE: The present study demonstrated that RJ is composed of muscarinic receptor agonist(s), likely ACh, and induces vasorelaxation through nitric oxide (NO) production from the vascular endothelium of healthy rats, leading to increased tail blood circulation. Thus, fresh RJ may improve peripheral circulation in healthy individuals.

Changes of myocardial lipidomics profiling in a rat model of diabetic cardiomyopathy using UPLC/Q-TOF/MS analysis.

BACKGROUND: Diabetic cardiomyopathy (DCM) is a serious cardiac dysfunction induced by changes in the structure and contractility of the myocardium that are initiated in part by alterations in energy substrates. The underlying mechanisms of DCM are still under controversial. The observation of lipids, especially lipidomics profiling, can provide an insight into the know the biomarkers of DCM. The aim of our research was to detect changes of myocardial lipidomics profiling in a rat model of diabetic cardiomyopathy. METHODS: Diabetic cardiomyopathy was induced by feeding a high-sucrose/fat diet (HSFD) for 28 weeks and streptozotocin (30 mg/kg, intraperitoneally). The ultra-high-performance liquid chromatography (UPLC) coupled to quadruple time-of flight (QTOF) mass spectrometer was used to acquire and analyze the lipidomics profiling of myocardial tissue. Meanwhile, parameters of cardiac function were collected using cardiac catheterization, and the cardiac index was calculated, and fasting blood glucose and lipid levels were measured by an ultraviolet spectrophotometric method. RESULTS: We detected 3023 positive ion peaks and 300 negative ion peaks. Levels of phosphatidylcholine (PC) (22:6/18:2), PC (22:6/18:1), PC (20:4/16:1), PC (16:1/18:3), phosphatidylethanolamine (PE) (20:4/18:2), and PE (20:4/16:0) were down-regulated, and PC (20:2/18:2), PC (18:0/16:0), and PC (20:4/18:0) were up-regulated in DCM model rats, when compared with control rats. Cardiac functions signed as values of left ventricular systolic pressure, maximal uprising velocity of left ventricular pressure and maximal decreasing velocity of left ventricular pressure were injured by 21-44%, and the cardiac index was increased by 25%, and fasting blood glucose and lipids were increased by 34-368%. Meanwhile, the cardiac lipid-related biomarkers have significant correlation with changes of cardiac function and cardiac index. CONCLUSIONS: UPLC/Q-TOF/MS analysis data suggested changes of some potential lipid biomarkers in the development of cardiac dysfunction and hypertrophy of diabetic cardiomyopathy, which may serve as potential important targets for clinical diagnosis and therapeutic intervention of DCM in the future.