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Deepti Varma & Dr. Susan Jansen, Temple University, Philadelphia, PA and David J. Tognarelli, Jasco Inc., Easton, MD Rapid Profiling of
Human Sex Hormones
Rapid separation of human sex
hormones in Biological Samples using
the JASCO X-LC system
Introduction:
Hypertension is a serious problem among African
breast cancer risk 3. Testosterone is a steroid hormone American population, especially women.
from the androgen group which is present at much According to the American Heart Association higher concentration in men than in women.
(AHA), around 42.6% of African American menabove the age 20 and 46.6% of African Americanwomen have hypertension1. According to another Figure 1: Conversion of Progesterone to Estrogenic
study conducted by AHA, hypertension was the Compounds
cause of death of 5,762 African American menand 6,664 African American women in 2004.1Men and postmenopausal women are known to havegreater incidences of hypertension compared to premenopausal women. This suggests that thefemale sex hormones have an effect on bloodpressure2. Estrone (E1), estradiol (E2), estriol (E3) and progesterone are the main female sexhormones in the body. Out of all these, estrone is assumed to be the only one present in variablequantities in postmenopausal women. 2- hydroxyesterone (2-OHE1), 16_-hydroxyestrone(16_ -OHE1) and 4- steroid hormone from the androgen group which is present at much higherconcentration in men than in women.
Determining the link between sex hormones withthe pathogenesis of hypertension is complex.
This application note describes a 6.1 min methodfor identifying a mixture of 8 sex hormones using Figure 2: Metabolism of Estrogenic Compounds
Jasco X-LC. Separation of these compounds using conventional HPLC typically takes around 16 minutes. hydroxyestrone (4-OHE1) are the majormetabolites of estrone. The ratio of 2- hydroxyesterone (2-OHE1) to 16_-hydroxyestrone (16_ -OHE1) is considered to be an index ofbreast cancer risk 3. HPLC typically takes around 16 minutes. hydroxyestrone (4-OHE1) are the major metabolites of estrone. The ratio of 2-hydroxyesterone (2-OHE1) to 16_-hydroxyestrone(16_ -OHE1) is considered to be an index of Experimental:
Results and Discussion:
Sample Preparation:
Figure 2 is an X-LC chromatogram of a standard All the hormones were dissolved in 1 ml methanol mixture of hormones (50 ng each). The compounds such that the stock mass concentration for all was are separated by a gradient elution of water and 1mg/ml.Working standards for the analytes were acetonitrile on a Restek Pinnacle DB Biphenyl 1.9_m prepared by serial dilution using ACN. 2-methoxy (50 x 2.1 mm) at a flow rate of 0.165 ml/min.
Estrone, 4-hydroxy estrone were obtained from Wavelength 220 nm was used for the detection of these compounds. Resolution of testosterone from hydroxyestrone were obtained from Steraloids, estrone, is 1.25 in this method, acceptable for estrone, E1; estradiol,E2; estriol, E3; were quantitative determinations of both species.
Figure 2.
X-LC chromatogram of a standard
XLC system and operating conditions:
mixture of hormones.(critical pair
resolution, 1.25)
Mobile Phase A: WaterMobile Phase B: ACNFlow Rate: 0.165 ml/min Time (min)
%B
References:
Stationary Phase:
1. American Heart Association- Statistical Fact S Column Temperature:
2. Khalil AR, Hypertension, 2005, 46, 249-254.
UV Detection:
3. Ursin G et al, Environ Health Perspect, 1997, 105, Injection Volume: 1_l
Contact:
Jasco, Inc.
28600 Mary’s Court, Easton, MD 21601
Phone: 800-333-5272 Fax: 410-822-7526
www.jascoinc.com

Source: http://www.jasco.com.br/imagem/catalogos/aplicacoes/cromatografia/xlc_07.pdf