Enzymic conversion of 3beta-hydroxy-5-ene-steroids and their sulfates to 3-oxo-4-ene-steroids for increasing sensitivity in LC-APCI-MS.
A method of increasing the sensitivity of 3beta-hydroxy-5-ene (Delta5)-steroids in liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) based on the structural conversion by cholesterol oxidase (ChO) was demonstrated. The Delta5-steroids were rapidly converted to their 3-oxo-4-ene (Delta4)-forms by the treatment with ChO and the obtained Delta4-forms provided 3-14-fold higher sensitivity compared to intact steroids in the positive-APCI-MS. This enzymic conversion method was also applied to the sulfated conjugates of Delta5-steroids after solvolysis. The method enabled the detection of trace levels of dehydroepiandrosterone and androstenediol 3-sulfate in human serum, which could not be detected by the usual LC-MS.
Higashi
Tatsuya
T
Division of Pharmaceutical Sciences, Graduate School of Natural Science and Technology, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Japan. higashi@p.kanazawa-u.ac.jp
Takayama
Naoki
N
Shimada
Kazutake
K
D000735:Androstenedione / Q000032:analysis / Q000737:chemistry*; D002626:Chemistry, Pharmaceutical / Q000379:methods; D002789:Cholesterol Oxidase / Q000032:analysis* / Q000737:chemistry; D002853:Chromatography, Liquid / Q000379:methods*; D003687:Dehydroepiandrosterone / Q000032:analysis / Q000737:chemistry*; D004345:Drug Industry / Q000379:methods; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D011284:Pregnenolone / Q000032:analysis / Q000737:chemistry*; D011374:Progesterone / Q000032:analysis / Q000737:chemistry*; D012680:Sensitivity and Specificity; D013256:Steroids / Q000032:analysis / Q000737:chemistry*; D013431:Sulfates / Q000737:chemistry*; D013739:Testosterone / Q000032:analysis / Q000737:chemistry*; D013997:Time Factors
D013256:Steroids; D013431:Sulfates; D013739:Testosterone; D000735:Androstenedione; D003687:Dehydroepiandrosterone; D011374:Progesterone; D011284:Pregnenolone; D002789:Cholesterol Oxidase