Determination of isosinensetin in rat plasma by UHPLC-MS/MS: Application to oral and intravenous pharmacokinetic study in healthy rats.

Isosinensetin is a polymethoxyflavone existing in various kinds of citrus. It has exhibited significant anti-proliferative activity and herb-drug interaction. To date, a specific determination method to quantify isosinensetin concentration in biological matrix has not been developed. In the present study, a highly specific, simple and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) approach was developed and validated for quantification of isosinensetin in rat plasma with subsequent application to a pharmacokinetic study. Isosinensetin and lysionotin (internal standard, IS) were extracted from rat plasma by a single step protein precipitation using acetonitrile as precipitation agent. The chromatographic separation was conducted using an Agilent C18 column with a gradient elution system (0.1 % formic acid aqueous solution and acetonitrile) within 3.5 min. An electrospray ionization (ESI) source operating in positive mode and multiple reaction monitoring (MRM) were used to monitor the transitions of m/z 373.1 → 343.1 for isosinensetin and m/z 345.1 → 315.1 for IS. The developed method was linear within the range of 1-1000 ng/mL and fully validated according to FDA guidelines. The accuracy values reported as relative errors were between 2.0 and 10.0 % for three quality control levels (2, 400 and 800 ng/mL) and lower limit of quantification (LLOQ). The precisions were ≤11.1 % for quality controls and ≤18.1 % for LLOQ. The recoveries and matrix effects of isosinensetin were in the range of 83.4-87.7 % and 105.6-108.8 %, respectively. Other parameters such as selectivity, carryover effect, dilution integrity and stability were also validated and met the acceptance criteria. The method was applied to a pharmacokinetic study in rats following oral and intravenous administration of isosinensetin. Isosinensetin was rapidly absorbed with a poor bioavailability of 2.19 % and quickly eliminated with mean half-life of 1.40 h and 1.76 h for oral and intravenous route, respectively.

UHPLC-MS/MS method for the quantification of aloin-A in rat plasma and its application to a pharmacokinetic study.

Aloin-A (also known as barbaloin), the main bioactive anthraquinone-C-glycoside of Aloe species, exhibits various beneficial pharmacological effects. However, the determination and pharmacokinetic study of aloin-A in rat plasma need to be improved and systematically demonstrated. In the present study, a simple, robust and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for rapid quantification of aloin-A in rat plasma was developed. Plasma preparation was conducted by a single step protein precipitation with obtusin serving as an internal standards (IS) followed by separation of the analytes using an Agilent C18 column with a gradient mobile phase comprised of acetonitrile and formic acid aqueous solution. Negative ion electrospray was used and multiple reaction monitoring transitions were m/z 417.1 → 297.0 for aloin-A and m/z 343.1 → 328.1 for IS, respectively. The developed method was validated with linear range of 1-1000 ng/mL. All validation parameters were well within the acceptance criteria based on the guidance of FDA. The validated approach was successfully applied to analyze samples from a pharmacokinetic study in healthy rats following intravenous and oral administration. Aloin-A was found to be quickly absorbed, extensively distributed and rapidly eliminated. The absolute bioavailability of aloin-A was 5.79%.

Baicalin alleviates hyperglycemia-induced endothelial impairment 1 via Nrf2.

Baicalin is the major component found in Scutellaria baicalensis root, a widely used herb in traditional Chinese medicine, which exhibits strong anti-inflammatory, anti-viral and anti-tumor activities. The present work was devoted to elucidate the molecular and cellular mechanisms underlying the protective effects of Baicalin against diabetes-induced oxidative damage, inflammation and endothelial dysfunction. Diabetic mice, induced by streptozotocin (STZ), were treated with intraperitoneal Baicalin injections. Human umbilical vein endothelial cells (HUVECs) were cultured either in normal glucose (NG, 5.5 mM) or high glucose (HG, 33 mM) medium in the presence or absence of Baicalin for 72 h. We observed an obvious inhibition of hyperglycemia-triggered oxidative damage and inflammation in HUVECs and diabetic aortal vasculature by Baicalin, along with restoration of hyperglycemia-impaired nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway activity. However, the protective effects of Baicalin almost completely abolished in HUVECs transduced with shRNA against Nrf2, but not with nonsense shRNA. Mechanistic studies demonstrated that HG decreased Akt and GSK3B phosphorylation, restrained nuclear export of Fyn and nuclear localization of Nrf2, blunted Nrf2 downstream target genes, and subsequently induced oxidative stress in HUVECs. However, those destructive cascade, were well prevented by Baicalin in HUVECs. Furthermore, LY294002 and ML385 (inhibitor of PI3K and Nrf2) attenuated Baicalin mediated Nrf2 activation and the ability of facilitates angiogenesis in vivo and ex vivo. Taken together, the endothelial protective effect of Baicalin under hyperglycemia condition could be partly attributed to its role in downregulating reactive oxygen species (ROS) and inflammation via the Akt/GSK3B/Fyn-mediated Nrf2 activation.

[Two new flavonoid glycosides from leaves of Moringa oleifera].

Two new flavonoid glycosides, quercetin-3-O-(4-O-crotonyl)-ß-D-glucopyranoside (1) and quercetin-3-O-[6-O-(2E)-pentenoyl]-ß-D-glucopyranoside (2), along with nine known ones, isoquercetin (3), astragalin (4), quercetin-3-O-(6-O-acetyl)-ß-D-glucopyranoside (5), kaempferol-3-O-(6-O-acetyl)-ß-D-glucopyranoside (6), quercetin-3-O-(6-O-crotonyl)-ß-D-glucopyranoside (7), kaempferol-3-O-(6-O-crotonyl)-ß-D-glucopyranoside (8), vitexin (9), isovitexin (10), and isorhamnetin-3-O-ß-D-glucopyranoside (11), were isolated from the leaves of Moringa oleifera by various chromatographic technologies. Their structures were elucidated by spectroscopic methods including UV, IR, MS, and NMR. In addition, compounds 7 and 8 were isolated from this plant for the first time.

Quantification and pharmacokinetics of alpinetin in rat plasma by UHPLC-MS/MS using protein precipitation coupled with dilution approach to eliminate matrix effects.

Alpinetin, a bioactive flavonoid, has attracted great attention due to its diverse therapeutic effects, namely anti-oxidant, anti-tumor and anti-inflammatory effects with low systemic toxicity. Various determination methods have been developed in quality control and plant chemistry areas. However, quantification and pharmacokinetics of alpinetin in biological matrix have not been studied. In the present research, a sensitive, efficient and reliable ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for the determination of alpinetin in rat plasma was developed and validated. Plasma samples were processed with protein precipitation (PP) followed by a 5-fold acetonitrile/water (50:50, v/v) dilution to significantly decrease matrix effect which exited in one step PP method. Determination of alpinetin was conducted using positive electrospray ionization tandem mass spectrometry in multiple reaction monitoring mode. Results demonstrated that the method was precise (3.3%-12.3%), accurate (-5.8% to 10.8%) and linear in the range of 1-1000 ng/mL. The new developed method was subsequently applied to a pharmacokinetic research of alpinetin following oral and intravenous dosing to healthy Sprague-Dawley rats. Alpinetin was demonstrated rapid absorption after oral administration with an absolute bioavailability of ∼15.1% and extensive distribution after dosing.