| To develop a RP-HPLC-FL method for determination of three bisbenzylisoquinoline alkaloids in the embryo of Nelumbo nucifera Gaertn. and compare it with RP-HPLC-UV. A Hypersil ODS-C18 column was used with the mobile phase of methanol-acetonitrile-water-triethylamine (20/40/40/0.12, v/v/v/v) at the detection wavelength ofλmax with the UV detector andλex +λem with the fluorescence detector. The flow rate was 1.0 ml/min. The fluorescence excitation and emission spectrum of the three alkaloids are similar.λex=280 nm andλem =620 nm. The linear range was 1-50μg/mL in the HPLC with the UV detector and 10-100μg/mL in the HPLC with the fluorensence detector. The RSD of inter-day and intra-day was 1.0-8.0% in both detectors. The recovery of three alkaloids were 95~105%. The LOD (lower limit of detection) of the four alkaloids was 250-500 ng/ml and LOQ (lower limit of quantification) was 500-1000 ng/ml with the UV detector. The LOD of them was 500-1000 ng/ml and LOQ was 1000-2500 ng/ml with the fluoresecene detector. Different groups of samples were determined by this two methods. The determination results of samples are similar. The HPLC-FL method is new, simple, reliable and precise for the determination of three alkaloids in the embryo of Lotus plumule.Part II: Effects of Neferine on the Pharmacokinetics of Amiodarone in Rats1. Improved RP-HPLC method for the determination of amiodarone in small volumes of plasmaExisting methods to determine amiodarone either with the use of a large volumes of plasma or with the large cost of money in solid phase extraction. In this study, we showed that a simple and sensitive HPLC-UV method to determine the concentration of amiodarone in small volumes of rat plasma. The chromatographic conditions were ODS RP-C18 column (4.6×200 mm), mobile phase methanol-water-acetonitrile-glacial acetic acid (200/36/23.6/8.6, v/v; pH 7.53), and flow rate 1.2 ml/min, detection wavelength 242 nm: 1, I.S.; 2, amiodarone; 3, possible metabolites of amiodarone. Retention times for amiodarone and I.S. were approximately 7.5 and 19.5 min, respectively. In this study, we conjoined use the liquid-liquid extraction and the protein precipitation. Extraction conditions were optimized by using several solvents such as diethyl ether, acetoacetate, dichloromethane, methanol, acetonitrile and combinations of them. Among these, dichloromethane gave the highest recovery and least interference at the area of amiodarone and I.S.. The extraction recoveries of amiodarone (n=6) from spiked rat plasma were satisfactory at low, middle and high concentrations, which varied from 96.76% to 102.22%, which were consistent with SFDA guidelines. Recovery of the I.S. of tamoxifen was very consistent with a mean of 89.86% and a R.S.D. of 4.8% (n=6). The intra-day precision (expressed as percent relative standard deviation, R.S.D.%) ranged from 4.9 to 9.4% and the intra-day accuracy (expressed as percent of nominal values) ranged from 97.8 to 98.9%. The method showed reproducibility with inter-day precision ranging from 7.9 to 10.1%. The inter-day accuracy ranged from 94.3 to 102.7%. These results indicated that the present method has a satisfactory accuracy, precision and reproducibility. Stock solutions of amiodarone and I.S. were stable when stored at room temperature for 6 h and after storage at 4 oC for 5weeks. Table 4 displays stability for amiodarone under the following conditions: (1) stability of amiodarone in rat plasma through three freeze-thaw cycles; (2) short-term stability of amiodarone in rat plasma at room temperature for 6 h; (3) long-term stability of amiodarone stored at -70 oC for 1 month; (4) the post-preparative stability of amiodarone during storage at 4 oC for 12 h. 2. Effects of Neferine on the Pharmacokinetics of Amiodarone in RatsAmiodarone is an iodinated benzofuran derivative with predominantly Class III anti-arrhythmic effects and that is used to treat supraventricular and ventricular arrhythmias. The purpose of this study was to assess the potential of neferine, which is an effective anti-pulmonary fibrosis drug isolated from the embryo of traditional Chinese medicine and herbs-Nelumbo nucifera Gaertner's seeds, to alter the pharmacokinetic profile of amiodarone. Experimental Sprague-Dawley (SD) rats were randomly divided into 2 groups to evaluate the effects of neferine on the pharmacokinetics of amiodarone by oral and intravenous administration. Blood samples were collected from the orbital venous plexus at indicated time points and were analyzed for amiodarone using RP-HPLC. The geometric mean ratio for Cmax and AUC0-96 was calculated. The pharmacokinetics parameters of amiodarone intravenously administered or orally administered were not significantly different from the control (without neferine) group (with ratios of 0.7~1.4 in all experimental groups), suggesting that neferine had no effect on amiodarone plasma pharmacokinetics. The dosage regimen of amiodarone should not be taken into consideration for toxic reactions when combined with neferine. |
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