Simultaneous analysis of doping drugs in human plasma and urine using HPLC- DAD
AhighlysensitiveRP-HPLCmethod has been developedforsimultaneousseparation and quantitation ofseven doping drugs, includingfour diureticsu00ebHydrochlorothiazide (HCTZ), Furosemide (FUR), Indapamide (IDP) and spironolactone (SPIRO)u00ed, Salbutamol (SAL) as u201a-agonist, Testosterone (TSE) as anabolic and Betamethasone (BMS) as corticosteroid in spiked human plasma and urine, by usingZorbaxeclipseHC-C18column(250mmx4.6mmx5u00b5m)withmobilephase acetonitrile: phosphoric acid pH3 (50:50, v/v)underisocratic conditionswith flow rate of 1.0mlmin -1 and atroomtemperature.Diode array detectorwas adjusted at u00eb225, 272, 235,242 and 244u00ed and239nmforquantitativedetermination ofu00ebHCTZ, SAL, FUR,IDPandTSEu00edand both u00ebSPIROandBMSu00ed,respectively.The linearity range for the studied drugsin the plasma was 100-9000, 100-1800, 100-5000, 200-9000 and 1000-9000 ng.ml -1for u00ebHCTZ and SALu00ed,for u00ebFUR andTSEu00ed, for IDP,for SPIRO and forBMS,respectively. LODs and LOQs values were found to be u00eb31.16, 29.99, 28.14, 29.84, 31.98, 28.55 and 250.99u00ed and u00eb94.42, 90.88, 85.27, 90.42, 96.91, 86.52 and 760.58u00edngml -1 forHCTZ, SAL, FUR,IDP,TSE, SPIROandBMS,respectively.Also;theinvestigated drugs could bedetermined in spiked urinesamples after directdilutionandsolidphase extraction(SPE),where in the last way (SPE) HCTZ and SAL could not be determined, since they give irreproducible results.Indirectdilutionway;the linearityrangewas 150u00f1 5000, 50 u00f1 5000, 150 u00f1 1500 and 100 u00f1 5000 ng.ml -1 for u00ebHCTZ and BMSu00ed, u00ebSAL, IDP and TSEu00ed,u00ebFURu00ed and for u00ebSPIROu00ed,respectively and the u00ebLODs and LOQs-val- uesu00ed were u00eb39.41, 11.98, 35.52, 12.70, 14.11, 29.01 and 40.72u00ed and u00eb119.42, 36.30, 107.64, 38.48, 42.76, 87.91 and 123.39u00ed ng.ml -1 for HCTZ, SAL, FUR, IDP,TSE, SPIROandBMS,respectively.In SPEmethod;the linearity rangewas 250-3000, 150- 6000 and 150-7000 ng.ml -1 for FUR,IDP and foru00ebTSE, SPIRO and BMSu00ed, respectively and the u00ebLODs and LOQs-valuesu00edwere u00eb70.11, 39.15, 42.71, 45.91 and 49.01u00edand 212.45, 118.64, 129.42, 139.12 and 148.52 ng.ml - 1 for FUR, IDP, TSE, SPIRO and BMS, respectively. It wasshown that SPE is more sensitive,for determination ofFUR,IDP,TSE, SPIROandBMS, than direct dilution(only1:4dilutioncomparedto1:50folddilutionindirectdilution),however, HCTZandSALcould not be determined.
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