西咪替丁是一种治疗消化性溃疡的药物。为了分析其生物利用度需要分析其血清样品的浓度。在全血,尿液或者血清分析中,长需要大量的手动固相萃取对样品进行进化。
此篇报道叙述了使用在线固相萃取-液相联用直接分析生物样品的过程。整套分析设备由溶剂输送系统,自动进样器,和在线固相萃取控制系统组成。只需手动用溶剂将血清样品进行1:1的稀释后,就能直接进行分析。
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.
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