ABSTRACT
Objective::To study the pharmacokinetics of sinapic acid from stir-fried Raphani Semen in normal rats and the correlation between pharmacokinetics-pharmacodynamics (PK-PD) in asthma rats. Method::Normal rats received 4.5, 9, 18 g·kg-1 of stir-fried Raphani Semen by oral administration, respectively. Blood was taken from ophthalmic venous plexus at different time points according to the experimental design, the plasma concentration of sinapic acid was analyzed by UHPLC-MS/MS, and data analysis was performed using DAS 3.2.8 software to obtain the pharmacokinetic parameters. Rat asthma model was established by intraperitoneal injection of ovalbumin with aluminum hydroxide, and treated with ethanol extract of stir-fried Raphani Semen (low and high doses of 4.5, 9 g·kg-1). After treatment for 3 weeks, taking blood at different time points, plasma and serum were separated. UHPLC-MS/MS was established for the determination of plasma concentration of sinapic acid, contents of interleukin-5 (IL-5), immunoglobuin E (IgE), tumor necrosis factor-α (TNF-α) in serum at different time points were detected by enzyme-linked immunosorbent assay (ELISA), DAS 3.2.8 software was used for PK-PD model fitting and data analysis. Result::After normal rats were administrated with low, medium and high doses of stir-fried Raphani Semen, the peak concentration (Cmax) of sinapic acid in plasma were (29.35±10.32), (62.70±27.47), (137.33±40.95) μg·L-1, its area under the curve (AUC0-t) were (92.83±27.16), (240.74±75.09), (633.95±195.88) μg·L-1·h, its peak time (Tmax) were (2.58±0.80), (3.00±0), (5.50±1.23) h, respectively. Compared with the low dose group, AUC0-t and mean retention time (MRT0-t) were all increased in the medium and high dose groups, showing statistical differences (P<0.05, P<0.01). The linear relationship of AUC0-t in sinapic acid was good within the dose range of 4.5-18 g·kg-1. After treating with ethanol extract of stir-fried Raphani Semen for 0.083, 0.167 h, compared with the model group of asthmatic rats, serum levels of IL-5, IgE, TNF-α of the medication groups were decreased to different degrees (P<0.05, P<0.01). Cmax of sinapic acid in the low and high dose groups were (58.43±29.94), (61.16±18.79) μg·L-1, its AUC0-t were (188.75±37.07), (247.90±36.89) μg·L-1·h, respectively. AUC0-t, apparent volume of distribution (Vz/F) and clearance rate (CLz/F) all increased significantly with the increase of dose. The best pharmacokinetic model of sinapic acid was fitted as a one-compartment model for extravascular administration, PK-PD model may be applicable to indirect connection model. Conclusion::The plasma concentration of sinapic acid is correlated with contents of IL-5, IgE and TNF-α, dosage and functional state (pathological or physiological state) can affect the pharmacokinetic behavior of sinapic acid from stir-fried Raphani Semen in rats, and it has a certain correlation with the anti-asthmatic effect.
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Objective: To analyze the dynamic changes of components in the enzymolysis process of raw products of Raphani Semen. Method: HPLC was employed to analysis of characteristic spectra of Raphani Semen at different enzymolysis time with mobile phase of acetonitrile-0.1% phosphoric acid aqueous solution for gradient elution and detection wavelength at 225 nm.The characteristic peaks were calibrated, meanwhile, the UV spectra of characterstic peaks were extracted, and the difference between UV spectra and the changes of peak areas were compared, and the dynamic changes of characterstic components in Raphani Semen were analyzed. Result: Eleven characteristic peaks were marked from the characteric spectra of raw products of Raphani Semen at different enzymolysis time, and glucoraphenin and sinapine thiocyanate were assigned.Glucoraphenin was enzymatically hydrolyzed fastly by myrosinase, and an intermediate was generated, and then continue to be decomposed into other components.Sinapine thiocyanate did not change significantly during the enzymolysis process, and sinadiosides was also enzymatically degraded. Conclusion: The enzymolysis of Raphani Semen is not only the glucoraphenin, but also the sinadiosides.This paper can provide reference for the property change of Raphani Semen in processing.
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Objective: To explain the phenomenon that Panax ginseng is not compatible with Raphani Semen based on pharmacodynamics and pharmacokinetics. Methods: The forced swimming time and biochemical parameters such as blood lactate (BLA), serum urea nitrogen (SUN), and hepatic glycogen (GLU) were determined for anti-fatigue experiment. The UPLC-MS/MS was used to analyze the pharmacokinetics of Rg1, Re, Rb1, and Rd after orally administration of P. ginseng and P. ginseng combined with Raphani Semen to rats. Pharmacokinetic differences of four ginsenosides between single uses of P. ginseng and combined with Raphani Semen were investigated. Results: The results showed that Raphani Semen tended to significantly reduce the anti-fatigue activity of P. ginseng. Co-administration of P. ginseng and Raphani Semen had significant effects on the pharmacokinetics of the four ginsenosides in rats compared to that observed with P. ginseng extract alone. The AUC0–12 h values of the four ginsenosides in PG group were higher than the corresponding values in the PR group. It can be inferred that Raphani Semen decreased the blood exposure of the four ginsenosides in rats when it combined with P. ginseng. Conclusion: The anti-fatigue activity and pharmacokinetic results showed that Raphani Semen may reduce the pharmacological actions of P. ginseng.
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To establish an accurate, rapid and efficient method for authenticating Cuscutae Semen and Raphani Semen by using rapid PCR amplification. The samples of Cuscutae Semen, Raphani Semen and their adulterants were collected. The total DNA of the samples has been extracted, and ITS sequence from Cuscutae Semen, Raphani Semen and their adulterants was amplified by PCR and sequenced directionally. These sequences were aligned by using Clustal W. Specific primers were designed and amplified by two-steps PCR amplification method. The rapid PCR methods for authenticating Cuscutae Semen and Raphani Semen were established by optimizing the denatured and annealing temperature, cycle numbers, and etc. When 100 × SYBR Green I was added in the PCR product, strong green fluorescence was visualized under 365 nm UV lamp whereas adulterants showed no florescence. The results indicated that the rapid PCR method can identify Cuscutae Semen and Raphani Semen rapidly. This study provides the technical support for authentication of Chinese medicinal materials.