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Determination of the contents of 6β-hydroxycortisol and hydrocortisone in healthy human's urine by RP-HPLC

WAN Jun,ZHANG Xiaoguang,CHEN Jinghai,CHEN Yao(Department of Pharmacy,Xiamen Maternity and Child Care Hospital,Xiamen 361003,China)  
AIM To establish a method for determining the concentration of 6β-hydroxycortisol and hydrocortisone in human urine.METHODS The RP-HPLC separation using gradient elution was performed on a SHIMADZU Insertil ODS-3(250 mm×4.6 mm,5μm))reversion phase chromatography column at column temperature of 30℃.The mobile phase consisted of water-acetonitrile,and the detection wave-length was at 240 ran.The urine was extracted with liquidphase extraction method by ethyl acetate.RESULTS The linear regress equations for 6β-hydroxycortisol and hydrocortisone were y=0.004 1x-0.119 2,r=0.999 0 and y=0.012 7x-0.050 5,r=0.997 8,respectively.The percentages of their average recoveries were both above 95%,and the intra-day RSD and inter-day RSD were less than 5%. The concentration of 6β-hydroxycortisol and hydrocortisone in 20 volunteers' urine was(151.90±94.05)μg·L~(-1)and (26.38±20.07)μg?L~(-1)respectively and the ratio was 6.32±2.36.CONCLUSION The present method is highly specific,sensitive,simple and reliable,and can be used for the large sample analysis.
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