化学药中主要物质含量分析检测方案

Analysis of Cimetidine in plasma using On-line solid phaseextraction to HPLC ProspektClinical June 1996 Introduction Cimetidine (Tagamet) is an H receptor antagonistcommonly taken orally to fight peptic ulcers.To evaluate clinical efficacy and bioavailability ofCimetidine， plasma samples from pharmacokineticstudies are analysed to establish the data required.Many manual solid phase extraction methods exist forCimetidine in whole blood， urine and plasma， etc.The following method describes the automation of theSPE method followed by On-line HPLC analysis. Experimental Figure 1： System switching diagram Instrumentation For chromatographic separations a Varian Star 9012 HPLC pump， Star 9050 UV-Vis detector and Starworkstation were used. For the solid phase extractionprocedure a Solvent Delivery Unit (SDU) was used to condition and to purge thecartridges with various solvents， aMARATHON autosampler to inject thediluted plasma samples (0.5 ml) into thesolvent stream and the Spark HollandPROSPEKT system fo control the On-lineextraction and pre-concentration ondisposable SPE cartridges. Chromatographic conditions Anal. Column： Varian ResElut C18， 4.6 mm × 15 cm SPE cartridge： C8 Analytichem (40-63 um)，10 ×2 mm， Spark Holland. Mobile phase： 0.01 M Ammonium Acetate/ Acetonitrile (4：1 v/v) Flowrate： 1.5 ml/min Detection： UV 228 nm Sample preparation Plasma samples are diluted with 0.01 M ammoniumAcetate in the ratio 1：1 prior to extraction. Afterwardsthe following sample preparation program was used： 1..Change cartridge. 2. Activate the cartridge with 1.0 ml methanol at1.5 ml/min. 3. Condition the cartridge with 2 ml 0.01 MAmmonium Acetate at 1.5 ml/min. 4. Load 0.5 ml diluted plasma sample with 3 ml0.01 M Ammonium Acetate at 1.5 ml/min. 5. Elute the Cimetidine and internal standardfrom the SPE cartridge onto the analyticalcolumn by the HPLC mobile phase during20 seconds. 6. End program. S.D.U. Marathon Prospekt Time Solv Purge flow Inj.valve Change Cart. Valve I Aux ENDTIME Comment MM：SS ml/min FI ： LoadF2： Inject F1Execute F1 ： EluleF2 ： Purg 0：00 1.5 F1 F2 Activate 0：40 2 Condifion 2：00 F2 Extraction 4：00 1 F1 On Elution ofcartridge 4：01 Off Purge tubing 4：20 F2 5：00 0 (0 10：00 END End of program Solvents： 1：Methanol； 2：0.01 M Ammonium Acetate Solvents： 1：Methanol； 2：0.01 M Ammonium Acetate o nn 寸寸6‘N O寸N M寸N H A chromatogram of Cimetidine and internal standard at a level of 0.1 pg/ml plasma (Cimetidine) is shown in figure 2. The compounds are well separated and -18，557 H + _9.167 -0.971 Results and discussion Chromatographic separation {r OO ie e ww ON ONCD EU clean-up has been effective. O守 wo>anboE dEOOH 0E E的NN ni puE Dnpo Op oZODNU 寸6HTU d TH 寸NOZOO… ‘‘价 n Figure 2： Chromatogram of Cimetidine and Internal standard af 0.1 ug/ml in plasma ofter on-line SPE-HPLC. References Mary Burke and Brian MckennaElan Chemical Company，Athlone， Republic of lreland Kevin Fernandez， Phil Kilby， Varian Ltd.，28 Manor Road，Walton-on-Thames，Surrey KT12 2QF， England，Tel. (01932) 898000，Fax. (01932) 228769.