Identification of Sanguinarine Metabolites in Rats Using UPLC-Q-TOF-MS/MS.

Sanguinarine (SAN), as the main active component of a traditional Chinese veterinary medicine, has been widely used in the animal husbandry and breeding industry. However, the metabolites of SA are still uncertain. Therefore, this research aimed to investigate the metabolites of SA based on rats in vivo. The blood, feces, and urine of rats were collected after the oral administration of 40 mg/kg SAN. Ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS) was employed to identify the metabolites of SAN. The elemental composition of sanguinarine metabolites was inferred by analyzing their exact molecular weight, and the structures of the metabolites were predicted based on their fragment ions and cleavage pathways. A total of 12 metabolites were identified, including three metabolites in the plasma, four in the urine, and nine in the feces. According to the possible metabolic pathways deduced in this study, SAN was mainly metabolized through reduction, oxidation, demethylation, hydroxylation, and glucuronidation. This present research has summarized the metabolism of SAN in rats, which is helpful for further studying the metabolic mechanism of SAN in vivo and in vitro.

Determination of multicomponents from Zhuanggu Guanjie capsule in rat plasma by UHPLC-MS/MS and pharmacokinetic study.

Aim: To develop a UHPLC-MS/MS method for the quantification of 12 constituents in rat plasma after oral administration of Zhuanggu Guanjie. Methods: Constituent separation was performed on a C18 column, and the mass spectrometric detection was performed in multiple reaction monitoring mode with a positive-negative ionization mode. Results: The method was successfully applied to the pharmacokinetic study of these 12 constituents after rats taking Zhuanggu Guanjie capsules. The results showed that psoralen, isopsoralen and aspersaponin VI were the key effective components and had high exposure. Conclusion: A rapid, simple and sensitive ultra-high-performance liquid chromatography coupled with triple quadrupole mass spectrometry method for the detection of 12 components in rat plasma after taking Zhuanggu Guanjie was developed and applied in this pharmacokinetics study.

Metabolite profiles of Gansuibanxia decoction in rat plasma and urine by UHPLC-Q-TOF/MS analysis.

Gansuibanxia decoction (GSBXD), a traditional Chinese medicine (TCM) formula with 2000 years of history, has good efficacies on treating cancerous ascites, pleural effusion, etc. However, little is known about its metabolite profiles owing to the lack of in vivo studies. In this study, we explored the prototypes and metabolites of GSBXD in rat plasma and urine using UHPLC-Q-TOF/MS technology. A total of 82 GSBXD-related xenobiotic bioactive components (38 prototypes and 44 metabolites) were confirmed or tentatively characterized, including 32 prototypes and 29 metabolites in plasma, and 25 prototypes and 29 metabolites in urine. The results showed that the bioactive components absorbed in vivo mainly contained diterpenoids, triterpenoids, flavonoids and monoterpene glycosides. Both phase I reactions (methylation, reduction, demethylation, hydrolysis, hydroxylation and oxidation) and phase II reactions (glucuronidation and sulfation) were involved in the metabolism of GSBXD in vivo. This study will provide a foundation for the quality control, pharmacological study and clinical application of GSBXD.

High-Dose Spermidine Supplementation Does Not Increase Spermidine Levels in Blood Plasma and Saliva of Healthy Adults: A Randomized Placebo-Controlled Pharmacokinetic and Metabolomic Study.

(1) Background: Spermidine is a biogenic polyamine that plays a crucial role in mammalian metabolism. As spermidine levels decline with age, spermidine supplementation is suggested to prevent or delay age-related diseases. However, valid pharmacokinetic data regarding spermidine remains lacking. Therefore, for the first time, the present study investigated the pharmacokinetics of oral spermidine supplementation. (2) Methods: This study was designed as a randomized, placebo-controlled, triple-blinded, two-armed crossover trial with two 5-day intervention phases separated by a washout phase of 9 days. In 12 healthy volunteers, 15 mg/d of spermidine was administered orally, and blood and saliva samples were taken. Spermidine, spermine, and putrescine were quantified by liquid chromatography-mass spectrometry (LC-MS/MS). The plasma metabolome was investigated using nuclear magnetic resonance (NMR) metabolomics. (3) Results: Compared with a placebo, spermidine supplementation significantly increased spermine levels in the plasma, but it did not affect spermidine or putrescine levels. No effect on salivary polyamine concentrations was observed. (4) Conclusions: This study's results suggest that dietary spermidine is presystemically converted into spermine, which then enters systemic circulation. Presumably, the in vitro and clinical effects of spermidine are at least in part attributable to its metabolite, spermine. It is rather unlikely that spermidine supplements with doses <15 mg/d exert any short-term effects.

Multicomponent Characterization of the Flower Bud of Panax notoginseng and Its Metabolites in Rat Plasma by Ultra-High Performance Liquid Chromatography/Ion Mobility Quadrupole Time-of-Flight Mass Spectrometry.

The flower bud of Panax notoginseng (PNF) consumed as a tonic shows potential in the prevention and treatment of cardiovascular diseases. To identify the contained multi-components and, in particular, to clarify which components can be absorbed and what metabolites are transformed, unveiling the effective substances of PNF is of vital significance. A unique ultrahigh-performance liquid chromatography/ion mobility quadrupole time-of-flight mass spectrometry (UHPLC/IM-QTOF-MS) profiling approach and efficient data processing by the UNIFITM bioinformatics platform were employed to comprehensively identify the multi-components of PNF and the related metabolites in the plasma of rats after oral administration (at a dose of 3.6 g/kg). Two MS2 data acquisition modes operating in the negative electrospray ionization mode, involving high-definition MSE (HDMSE) and data-dependent acquisition (DDA), were utilized aimed to extend the coverage and simultaneously ensure the quality of the MS2 spectra. As a result, 219 components from PNF were identified or tentatively characterized, and 40 thereof could be absorbed. Moreover, 11 metabolites were characterized from the rat plasma. The metabolic pathways mainly included the phase I (deglycosylation and oxidation). To the best of our knowledge, this is the first report that systematically studies the in vivo metabolites of PNF, which can assist in better understanding its tonifying effects and benefit its further development.

Global characterization of chemical compounds in Shidan granule and their metabolites in rat plasma using ultra-high-performance liquid chromatography coupled with quadrupole-time of flight mass spectrometry.

Shidan granule is an in-hospital traditional Chinese medicine prescription with obvious clinical effects in the treatment of chronic atrophic gastritis. Characterizing the compounds of Shidan granule and exploring the absorbed prototype constituents and metabolites in the blood is significant to understand its effective compounds. In this work, a reliable approach based on ultra-high-performance liquid chromatography combined with quadrupole time-of-flight mass spectrometry was performed for systematic analysis of chemical substances of Shidan granule and their metabolites in rat plasma. The compounds were rapidly characterized using integrated filtering methods, including mass defect filtering, neutral loss filtering, and diagnostic fragment ions filtering. As a result, 87 compounds were tentatively or unambiguously characterized. In rat plasma, a total of 79 components, including 21 prototype constituents and 58 metabolites, were preliminarily determined. And flavonoids-related, phenolic acids-related, saponins-related and terpenoids-related compounds were the main precursors of these metabolites. Phase I reactions (methylation, demethylation, hydroxylation, hydrogenation, and oxidation) and phase II reactions (glucuronidation, glucosidation, and sulfation) were observed as the major metabolic pathways of Shidan granule. It is the first comprehensive study on the metabolism of Shidan granule in vivo, which could offer a solid scientific basis for exploring the multiple effects and therapeutic mechanisms of Shidan granule.

Comprehensive characterization of the chemical composition of Lurong dabu decoction and its absorbed prototypes and metabolites in rat plasma using UHPLC-Q Exactive Orbitrap-HRMS.

Lurong Dabu decoction (LRDBD) is an effective traditional Chinese Korean ethnic medicine prescription composed of eight herbs, which is used for treating asthma. However, its material basis has not been studied yet. Herein, the use of a new and highly sensitive UHPLC-Q Exactive Orbitrap-HRMS technique is proposed for the high-resolution and accurate identification of the material basis of LRDBD. We identified 122 compounds belonging to different groups in LRDBD. Among these, 23 ingredients produced by decoction were identified and compared with 8 single herb compounds. Moreover, 39 other significantly different compounds were identified. Additionally, 29 absorbed prototype components and 35 metabolites were identified in rat plasma. Half of the prototype components were originated from antler velvet, it has corroborated the compatibility theory of Sasang medicine. To the best of our knowledge, the material basis of LRDBD was characterized for the first time. Our findings provide basic data and a method for further discovering potential drug targets and revealing the action mechanism of LRDBD in asthma treatment.

Rapid characterization and identification of the chemical constituents and the metabolites of Du-zhi pill using UHPLC coupled with quadrupole time-of-flight mass spectrometry.

A reliable method using ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was established to conduct a comprehensive analysis of the chemical constituents of Du-zhi pill (DZP) as well as their metabolites in rat plasma, urine and feces after gastric perfusion. The efficient on-line mass data acquisition modes combined the various off-line mass data mining strategy was applied. A full mass scan was performed, and then accurate MS/MS datasets were obtained through the use of a multiple mass defect filter (MMDF) and dynamic background subtraction (DBS)-dependent data acquisition method. Furthermore, post-acquisition data processing was conducted using various data-mining tools, including extracted ion chromatography (XIC), mass defect filtering (MDF), product ion filtering (PIF), and neutral loss filtering (NLF) (MetabolitePilot™). Finaly, a total of 176 compounds were identified or tentatively characterized in DZP. Moreover, a total of 233 components in vivo, which includes 92 prototype components and 141 metabolites, were unambiguously or tentatively identified in rat plasma, urine and feces. The metabolic pathways, including phase I reactions (hydroxylation, dehydroxylation and hydrogenation) and phase II reactions (acetylation, sulfation, glucuronidation and methylation), for the absorbed constituents, were explored and summarized. This is the first systematic study on the components of DZP and their metabolites in vivo. This study provide a valid analytical strategy for the characterization of chemical compounds and metabolites of TCM formulas. Moreover, an integrative strategy was proposed for the characterization and identification of chemical constituents and metabolites for additional TCM prescriptions.

HRPIF data mining based on data-dependent/independent acquisition for Rhei Radix et Rhizoma metabolite screening in rats.

In traditional Chinese medicine (TCM), components with identical nuclei often share structural similarity, indicating the possibility of similar second-level mass spectrometry (MS/MS) fragments. High-resolution product-ion filter (HRPIF) technique can be utilized to identify metabolites, with similar fragments, in vivo. In principle, this technique applies to TCM; however, its application has been restricted due to the limitations of traditional MS/MS data acquisition. Therefore, a novel analysis strategy, based on data-dependent acquisition (DDA) and data-independent acquisition (DIA) datasets, has been developed for the determination of template product ions and efficient non-targeted identification of TCM-related components in vivo by HRPIF and background subtraction (BS). This DDA-DIA combination strategy, taking Rhei Radix et Rhizoma as a test case, identified 71 anthraquinone prototype components in vitro (36 of which were discovered for the first time), and 45 related components in vivo, confirming glucuronidation and sulfation as the main reactions. The developed strategy could rapidly identify TCM-related components in vivo with high sensitivity, indicating the immense importance of this novel HRPIF data mining technology in TCM analysis.

Identification of metabolites in plasma related to different biological activities of Panax ginseng and American ginseng.

RATIONALE: Panax ginseng (PG) and American ginseng (AMG) are both medicinal plants of the Panax genus in the Acanthopanax family. Although PG and AMG have similar components of ginsenosides, there are many differences of their bioactivities. In this study, the biochemical mechanisms of different bioactivities of PG and AMG were explored by researching the differential metabolites in plasma after administration of each of PG and AMG. METHODS: In order to explore the material basis of differential bioactivities, two groups of mice were administrated orally with PG and AMG, and the method of metabolomics was used to identify the differential metabolites in plasma. Then network pharmacology was used based on the differential metabolites. Afterward, the metabolite-target-pathway network of PG and AMG was constructed; thus the pathways related to different bioactivities were analyzed. RESULTS: Through principal component analysis and orthogonal projections to latent structures discriminant analysis, there were 10 differential metabolites identified in the PG group and 8 differential metabolites identified in the AMG group. Based on network pharmacology, the differential metabolites were classified and related to differential bioactivities of PG and AMG. In the PG group, there were 6 metabolites related to aphrodisiac effect and exciting the nervous system, and 5 metabolites associated with raised blood pressure. In the AMG group, 5 metabolites were classified as having the effect of inhibiting the nervous system, and 6 metabolites were related to antihypertensive effect. CONCLUSIONS: This study explored the material basis of the differential biological activities between PG and AMG, which is significant for the research of PG and AMG use and to promote human health.

Translating n-3 polyunsaturated fatty acid status from whole blood to plasma and red blood cells during pregnancy: Translating n-3 status across blood fractions in pregnancy.

Women with low n-3 (omega-3) status in pregnancy can reduce their risk of early preterm birth (<34 weeks' gestation) through n-3 long chain polyunsaturated fatty acid (LCPUFA) supplementation. As investigators measure fatty acid status in different blood fractions, equations are needed to compare results across studies. Similarly, derived cut-points for defining low and replete n-3 status are needed to assist clinical interpretation during early pregnancy. Our aims were to develop equations to convert the percentage of total n-3 fatty acids, EPA+DHA and DHA between whole blood, plasma and red blood cells (RBC), and to derive cut-points for defining low and replete total n-3 fatty acid status in plasma and RBC from those already established in whole blood. Using blood samples from 457 pregnant women in a multicentre randomised controlled trial, equations for these interconversions were developed using simple linear regression models. Measures of n-3 fatty acid status in whole blood and plasma were strongly related (R2 > 0.85), while more moderate relationships were observed between measures in whole blood and RBC (R2 0.55 - 0.71), or plasma and RBC (R2 0.55 - 0.63). Using the conversion equations, established cut-points for low and replete n-3 status in whole blood (<4.2% and >4.9% of total fatty acids) converted to <3.7% and >4.3% of plasma total fatty acids, and to <7.3% and >8.1% of RBC total fatty acids. Agreement to define low and replete n-3 status was better between whole blood and plasma, rather than between whole blood and RBC. Our data also show that total n-3 fatty acids in plasma and serum are interchangeable. We conclude that either whole blood or plasma total n-3 fatty acids can be used to define low status in pregnancy and identify women who will most benefit from n-3 LCPUFA supplementation to reduce their risk of early birth. Further research is needed to determine the clinical utility of other fatty acid measures in various blood lipid fractions.

[Determination of cobalt in plasma blood samples by the ICP-MS method after oral intake of dietary supplements containing low doses of cobalt].

Salts of inorganic cobalt (Со) prevent the degradation of the alpha subunit of the hypoxia-inducible factor (HIF), imitating the state of hypoxia in the body and increasing the production of the endogenous hormone erythropoietin (EPO), and are used as doping substances that increase blood oxygen capacity and endurance, which give competitive advantages in sports. Currently, a large number of dietary supplements, including Co-containing ones, are offered on free sale. Their uncontrolled intake can affect not only the professional career of athletes, but also their health, due to the fact that this trace element and its salts are the strongest inorganic poisons and carcinogens. Despite this, their availability on the pharmaceutical market, a noticeable effect of erythropoiesis stimulation and a convenient oral form of administration lead to the need for their detection in modern doping control. The purpose of this research was to develop an approach to differentiate cobalt from vitamin B12, present in the body in its natural state, from the intake of cobalt salts by quantifying and comparing blood levels of vitamin B12 and total cobalt. Methods. The study involved 9 healthy volunteers (women and men) aged 25 to 45 years, leading an active lifestyle. Three of them took 2500 µg/day of cobalamin for 20 days (comparison group), three - dietary supplement containing cobalt asparaginate (100 µg/day in terms of pure cobalt), and the rest - dietary supplements with cobalt sulfate heptahydrate (100 µg/day in terms of pure cobalt) (administration groups) at the same time after meals. Blood samples were taken at baseline and on days 5, 9, 14 and 20. The concentrations of total cobalt in blood plasma samples of volunteers were measured by inductively coupled plasma mass-spectrometry (ICP-MS), the levels of cobalamin were determined on a Cobas 6000 immunochemical analyzer using the Elecsys Vitamin B12 II Assay ELISA kits. Results. It was found that oral intake of of cobalamin at a therapeutic dose significantly exceeding the recommended daily intake (3 µg), there was a regular slight increase in the blood concentration of total cobalt (1.1 times). At the same time intake of dietary supplements containing cobalt in the form of sulfate or asparaginate (about 100 µg per day in terms of pure cobalt) was accompanied by 4-6.7 fold increase in the concentration of total cobalt while unchanged vitamin B12 plasma concentration was observed. The detection of such changes can reliably indicate the use of prohibited salts and, of course, will be in demand for anti-doping control. Conclusion. Long-term monitoring of vitamin B12 and total cobalt levels, similar to hematological module of the Athlete Biological Passport program, will unambiguously detect possible abuse of cobalt salts and can be an additional evidence of the presence of these doping substances to other analytical methods, such as a combination of liquid chromatography and ICP-MS (LC-ICP-MS).

Multiple catechols in human plasma after drinking caffeinated or decaffeinated coffee.

BACKGROUND: Coffee is one of the most frequently consumed beverages worldwide. Research on effects of coffee drinking has focused on caffeine; however, coffee contains myriad biochemicals that are chemically unrelated to caffeine, including 3,4-dihydroxyphenyl compounds (catechols) such as caffeic acid and dihydrocaffeic acid (DHCA). OBJECTIVE: This prospective within-subjects study examined effects of drinking caffeinated or decaffeinated coffee on plasma free (unconjugated) catechols measured by liquid chromatography with series electrochemical detection (LCED) after batch alumina extraction. To confirm coffee-related chromatographic peaks represented catechols, plasma was incubated with catechol-O-methyltransferase and S-adenosylmethionine before the alumina extraction; reductions in peak heights would identify catechols. METHODS: Ten healthy volunteers drank 2 cups each of caffeinated and decaffeinated coffee on separate days after fasting overnight. With subjects supine, blood was drawn through an intravenous catheter up to 240 min after coffee ingestion and the plasma assayed by alumina extraction followed by LCED. RESULTS: Within 15 min of drinking coffee of either type, >20 additional peaks were noted in chromatographs from the alumina eluates. Most of the coffee-related peaks corresponded to free catechols. Plasma levels of the catecholamines epinephrine and dopamine increased with both caffeinated and decaffeinated coffee. Levels of other endogenous catechols were unaffected. Plasma DHCA increased bi-phasically, in contrast with other coffee-related free catechols. INTERPRETATION: Drinking coffee-whether caffeinated or decaffeinated-results in the rapid appearance of numerous free catechols in the plasma. These might affect the disposition of circulating catecholamines. The bi-phasic increase in plasma DHCA is consistent with production by gut bacteria.

A study of the influence of lead pollution on the anticoagulant activity of Whitmania pigra based on pharmacodynamics and metabolomics research.

Whitmania pigra Whitman (leech, also called Shuizhi in China, abbreviated as SZ), which has been used as a traditional Chinese medicine in the treatment of blood stasis syndrome (BSS) for a long time, is vulnerable to lead pollution in aquaculture environments. SZ has good anticoagulant activity. However, there are few studies on the influence of lead pollution on it. Therefore, we carried out the following researches to explore the influence of lead pollution on the anticoagulant activity of SZ and its mechanism. Firstly, the acute blood stasis model of rats was established by subcutaneous injection of adrenaline hydrochloride and ice water bath. Then unpolluted SZ (UPS) and lead-polluted SZ (LPS) were extracted. Next, the blood stasis model rats were administrated by gavage and the rats in normal control (NC) group and blood stasis model (BM) group were given the same amount of normal saline. Finally, the blood of the rats was collected to detect the coagulation function and hemorheology indexes. The metabolomics of rat plasma was studied by ultra-high-performance liquid chromatography coupled with orbitrap mass spectrometry (UPLC-Orbitrap-MS) technology. Principal component analysis (PCA), orthogonal partial least squares discriminant analysis (OPLS-DA) and Hierarchical clustering analysis (HCA) were used to perform metabolomics analysis. MetPA analysis was used to search for related metabolic pathways. The results of coagulation function and hemorheology showed that lead pollution could decrease the anticoagulant activity of SZ. The OPLS-DA score plots indicated that the plasma metabolites of rats in LPS group were close to BM group, while UPS group tended to be close to NC group both in the positive and negative ion mode. Hierarchical cluster analysis (HCA) suggested that UPS group and NC group were clustered into a branch, while LPS group and BM group were clustered into a branch. To sum up, lead pollution will reduce the anticoagulant activity of SZ. And lead pollution reduces the anticoagulant activity of SZ probably by influencing the metabolic pathways such as sphingolipid metabolism, amino acid metabolism and energy metabolism in rats.

Chemicalome and metabolome profiling of Chai-Gui Decoction using an integrated strategy based on UHPLC-Q-TOF-MS/MS analysis.

Traditional Chinese medicine prescriptions are widely believed to exert therapeutic benefits via a multiple-component and multiple-target mode. The systemic profiling of their in vitro chemicalome and in vivo metabolome is of great importance for further understanding their clinical value. Herein, an integrated strategy using ultra-performance liquid chromatography coupled with quadruple time-of-flight mass spectrometry was proposed to profile the chemicalome and metabolome of Chai-Gui Decoction. Particularly, an approach combined mass defect filter, characteristic product ion filter, and neutral loss filter was adopted to identify metabolites in plasma, urine, bile, and feces by MetabolitePilot. Consequently, a total of 174 constituents were identified or tentatively characterized and 70 metabolites that related to 21 representative structural components were matched in rat biofluids. Among them, 19 prototypes and 7 metabolites that contributed to flavonoids, monoterpenes, and phenylpropanoids were detected distribution in brain, heart, kidney, liver, lung or spleen. This study provided a generally applicable approach to comprehensive investigation on chemicalome and metabolome of traditional Chinese medicine prescriptions, and offered reasonable guidelines for further screening of quality control indicators of Chai-Gui Decoction.

Effects of spray-dried animal plasma on growth performance, survival, feed utilization, immune responses, and resistance to Vibrio parahaemolyticus infection of Pacific white shrimp (Litopenaeus vannamei).

Spray-dried animal plasma (SDP) in feed for several animal species provides health benefits, but research about use of SDP in shrimp feed is very limited. The objectives of the present study were to investigate the effects of dietary SDP on growth performance, feed utilization, immune responses, and prevention of Vibrio parahaemolyticus infection in Pacific white shrimp (Litopenaeus vannamei). In Experiment 1, the post-larvae were divided into five groups (four tank/group and 80 shrimp/tank) and fed four times daily diets with porcine SDP at 0, 1.5, 3, 4.5, and 6% of the diet for 45 days. In Experiment 2, the surviving shrimp from Experiment 1 were redistributed into six groups: four SDP groups as in Experiment 1 plus the positive and negative controls (four tank/group and 30 shrimp/tank). They were then challenged with V. parahaemolyticus by immersion at 105 colony-forming units (CFU)/mL and were fed with the same diets for another 4 days. In Experiment 1, shrimp fed 4.5% or 6% SDP diets had significantly higher body weight, survival rate, and improved feed conversion ratio. The immune parameters (total hemocyte count and phagocytic, phenoloxidase, and superoxide dismutase activities) of the shrimp fed 3-6% SDP diets also showed significant enhancement compared to the control. In Experiment 2, the survival rates of the 3-6% SDP groups were significantly higher than the positive control at day 4 after the immersion challenge. Likewise, the histopathological study revealed milder signs of bacterial infection in the hepatopancreas of the 3-6% SDP groups compared to the challenged positive control and 1.5% SDP groups. In conclusion, shrimp fed diets with SDP, especially at 4.5-6% of the diet, showed significant improvement in overall health conditions and better resistance to V. parahaemolyticus infection.

Development and Validation of a UPLC-MS/MS Method for the Quantitative Determination and Pharmacokinetic Analysis of Cirsimarin in Rat Plasma.

Cirsimarin is a bioactive antilipogenic flavonoid isolated from the cotyledons of Abrus precatorius and represents one of the most abundant flavonoids present in this plant species. Cirsimarin exhibits excellent antioxidant, lipolysis, and other biological properties; it can effectively trigger lipid movement and demonstrates antiobesity effects. In this work, an ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed for the determination of cirsimarin in rat plasma after intravenous administration. A standard curve of cirsimarin in blank rat plasma was generated over the concentration range of 1-3000 ng/mL. Six rats were administered cirsimarin intravenously (1 mg/kg). The method only required 50 µL of plasma for sample preparation, and the plasma proteins were precipitated with acetonitrile to pretreat the plasma sample. The precisions of cirsimarin in rat plasma were less than 14%, while the accuracies varied between 92.5% and 107.3%. In addition, the matrix effect varied between 103.6% and 107.4%, while the recoveries were greater than 84.2%. This UPLC-MS/MS method was then applied in measuring the pharmacokinetics of cirsimarin in rats. The AUC(0-t) values of cirsimarin from the pharmacokinetic analysis were 1068.2 ± 359.2 ng/mL·h for intravenous administration. The half-life (t 1/2) was 1.1 ± 0.4 h (intravenous), indicating that the metabolism of the compound was quick in the rats. Exploring the pharmacokinetics of cirsimarin in vivo can help better understand its metabolism.

Characterization of Metabolites in Plasma, Urine and Feces of Healthy Participants after Taking Brahmi Essence for Twelve Weeks Using LC-ESI-QTOF-MS Metabolomic Approach.

Brahmi essence, developed from Bacopa monnieri (L.) Wettst. standardized extract and mulberry juice, was proven to improve the memory speed of healthy participants aged 55-80 years old, following a 12-week dietary program. However, the metabolites have not yet been reported. Our objective was to characterize the altered metabolites in the plasma, urine, and feces of healthy volunteers after consumption of Brahmi essence for 12 weeks, using the LC-MS metabolomics approach. The altered metabolites were selected from OPLS-DA S-plots; 15 metabolites in the plasma, 7 in the urine, and 17 in the feces samples were tentatively identified by comparison with an online database and literature. The metabolites in the plasma samples were in the classes of amino acids, acylcarnitine, and phospholipids. Benzeneactamide-4-O-sulphate and 3-hydroxyhippuric acid were found in urine samples. The metabolites in the class of amino acids, together with jujubogenin and pseudojujubogenin, were identified in the fecal samples. The aminoacyl-tRNA, aromatic amino acids, and branched-chain amino acid biosynthetic pathways were mainly related to the identified metabolites in all three samples. It could be implied that those metabolites and their pathways might be linked with the effect of Brahmi essence on memory speed.

Vortex-assisted dispersive solid phase microextraction using Fe3O4/FeOOH magnetic nanocomposites for high-performance thin-layer chromatographic determination of zolmitriptan in rabbit plasma samples.

In this work, a fast, versatile, and convenient dispersive solid-phase micro-extraction (DSPME) method is combined with a spectro-densitometric technique for the analysis of zolmitriptan (ZOLM) in biological fluids. Fe3O4/FeOOH magnetic nanocomposites (MNCs) were prepared by a co-precipitation method in aqueous solutions and utilized subsequently as a sorbent in DSPME. By coupling DSPME with high-performance thin-layer chromatography (HPTLC) with fluorescence detection, the preconcentration and determination of (ZOLM) in presence of metoclopramide (MET) and paracetamol (PARA), which are prescribed as an adjuvant therapy with ZOLM, was accomplished. Adsorption capability was assessed using both Langmuir and Freundlich adsorption isotherm models. The adsorption data was fitted to Langmuir adsorption isotherm model as reflected by high determination coefficient (R2 = 0.9944). Moreover, adsorption kinetics was assessed by pseudo-first and pseudo-second order kinetic models. The data was fitted to pseudo-second order kinetics, which proves that ZOLM interaction with the adsorbent is a chemisorption process. Surface complexation with MNCs was suggested to explain the pH dependence of ZOLM sorption. The key parameters of extraction and desorption steps (including pH, extraction time, sample volume, magnetic adsorbent amount, and desorption circumstances) were evaluated. Optimized conditions for solid phase microextraction of ZOLM were pH 2.9, 5.0 mg Fe3O4/FeOOH MNCs, 15 min vortex-assisted extraction time and 3 × 200 µL of methanol: 33% ammonia; 4:1 as eluent. The analysis was achieved using ACN: dichloromethane: 33% ammonia (22.5: 6.0: 1.5, v/v/v) as a mobile phase and the fluorescence detection was carried out at 223 nm. The proposed DSPME method was successfully applied for trace quantification of ZOLM in rabbits' plasma (n = 6) after oral administration with a linearity range of 50.0 - 400.0 ng mL-1 (R2 = 0.9931), a detection limit of 12.0 ng mL-1 and extraction recovery of 97.27-99.89% with an RSD < 2% (n = 9). Moreover, the selectivity of the proposed approach for analysis of ZOLM in the presence of MET and PARA is demonstrated.

Increased dietary availability of selenium in rainbow trout (Oncorhynchus mykiss) improves its plasma antioxidant capacity and resistance to infection with Piscirickettsia salmonis.

Salmonid Rickettsial Septicaemia (SRS), caused by Piscirickettsia salmonis, is the most important infectious disease in the Chilean salmon farming industry. An opportunity to control this disease is to use functional micronutrients to modulate host mechanisms of response to the infection. Since P. salmonis may affect the host antioxidant system in salmonids, particularly that dependent on selenium (Se), we hypothesized that fish's dietary selenium supplementation could improve the response to the bacterial infection. To address this, we defined a non-antibiotic, non-cytotoxic concentration of selenium to evaluate its effect on the response to in vitro infections of SHK-1 cells with P. salmonis. The results indicated that selenium supplementation reduced the cytopathic effect, intracellular bacterial load, and cellular mortality of SHK-1 by increasing the abundance and activity of host glutathione peroxidase. We then prepared diets supplemented with selenium up to 1, 5, and 10 mg/kg to feed juvenile trout for 8 weeks. At the end of this feeding period, we obtained their blood plasma and evaluated its ability to protect SHK-1 cells from infection with P. salmonis in ex vivo assays. These results recapitulated the observed ability of selenium to protect against infection with P. salmonis by increasing the concentration of selenium and the antioxidant capacity in fish's plasma. To the best of our knowledge, this is the first report of the protective capacity of selenium against P. salmonis infection in salmonids, becoming a potential effective host-directed dietary therapy for SRS and other infectious diseases in animals at a non-antibiotic concentration.