ABSTRACT
Abstract A specific, sensitive and robust LC-MS/MS method was developed and validated for the quantification of deoxyelephantopin in rat plasma using simvastatin as an internal standard as per regulatory guidelines of Bioanalytical Method Validation. Plasma sample was prepared through liquid-liquid extraction. Chromatographic separation was performed on an Agela-C18 analytical column (1.8 µm, 2.1 mm × 50 mm) with an isocratic mobile phase consisting of 0.05% formic acid (dissolved in acetonitrile) and water (55:45, v/v) at a flow rate of 0.5 ml/min. The column oven was maintained at 40 ºC and the injection volume was 4 µl. Elution of deoxyelephantopin and the internal standard occurred at 5.1 and 6.3 min, respectively. The total chromatographic run time was 7.5 min. A linear response function was constructed in the concentration range of 13.2-2640 ng/ml. The intra- and inter-day precision and accuracy were in the range of 1.4-14.8% and -11.7 to 14.1%, respectively. The validated LC-MS/MS was successfully applied to the pharmacokinetic study of deoxyelephantopin after intravenous injection of 1, 2 and 4 mg/kg and oral administration of 7.5, 15 and 30 mg/kg deoxyelephantopin in rats. After oral and intravenous administration, the C max and AUC values of deoxyelephantopin increased dose-dependently.
ABSTRACT
BACKGROUND: Cimicifuga racemosa is one of the herbs used for the treatment of climacteric syndrome, and it has been cited as an alternative therapy to estrogen. Apart from hectic fevers, dyspareunia and so on, dry mouth also increase significantly after menopause. It has not yet been reported whether C. racemosa has any impact on the sublingual gland, which may relate to dry mouth. In an attempt to determine this, we have compared the effects of estrogen and C. racemosa on the sublingual gland of ovariectomized rats. RESULTS: HE staining showed that the acinar cell area had contracted and that the intercellular spaces were broadened in the OVX (ovariectomized rats) group, while treatment with estradiol (E2) and iCR (isopropanolic extract of C. racemosa) improved these lesions. Transmission electron microscopy showed that rough endoplasmic reticulum expansion in mucous and serous acinar epithelial cells and apoptotic cells was more commonly seen in the OVX group than in the SHAM (sham-operated rats) group. Mitochondria and plasma membrane infolding lesions in the striated ducts were also observed. These lesions were alleviated by both treatments. It is of note that, in the OVX + iCR group, the volume of mitochondria in the striated duct was larger than in other groups. Immunohistochemical staining showed that the ratio of caspase-3 positive cells was significantly increased in the acinar cells of the OVX group compared with the SHAM group (p < 0.05); and the MA (mean absorbance) of caspase-3 in the striated ducts also increased (p < 0.05). Estradiol decreased the ratio of caspase-3 positive cells and the MA of caspase-3 in striated ducts significantly (p < 0.05). ICR also reduced the ratio of caspase-3 positive cells and the MA in the striated ducts (p < 0.05), but the reduction of the MA in striated ducts was inferior to that of the OVX + E2 group (p < 0.05). CONCLUSION: Both estradiol and iCR can inhibit subcellular structural damage, and down-regulate the expression of caspase-3 caused by ovariectomy, but their effects were not identical, suggesting that both drugs confer a protective effect on the sublingual gland of ovariectomized rats, but that the specific location and mechanism of action producing these effects were different.