Published:Journal of Chromatographic Science,
ISSN 0021-9665Volume
39, Number 9, September 2001, pp. 385-387
Reversed-Phase
Liquid Chromatographic Method for Estrogen Determination in Equine Biological
SamplesSamira
B. Lima
Departamento de Reprodução Animal, Faculdade de Medicina Veterinária
e Zootecnia, Universidade de São Paulo-USP, Av. Prof. Dr. Orlando Marques
de Paiva, 87, 05508-900, São Paulo, SP, Brazil Ieda T.N. Verreschi
Departamento de Endocrinologia, Escola Paulista de Medicina, UNIFESP, Rua Pedro
de Toledo, 781, 13° andar, 04039-032, São Paulo, SP, Brazil Luciane M. Ribeiro Neto
Toxikón Assessoria Toxicológica, Rua Salvador Correa, 346, 04109-070,
São Paulo, SP, Brazil
Equine unsaturated estrogens are the main components of brand
formulations indicated for hormonal replacement therapy in both hypogonadic
and postmenopausal women. These hormones are produced by the fetoplacental unit
during equine gestation. A method is described for the quantitative determination
of equilenin (EL), equilin (EQ), 17a-dihydroequilin (17dEQ), and estrone (E1)
in the plasma of a pregnant mare. Blood samples are obtained weekly during pregnancy
by jugular venipuncture using sodium ethylenediaminetetracetic as the anticoagulant.
For the quantitation of these estrogens, plasma is submitted to enzymatic hydrolysis
followed by liquid–liquid extraction. A high-performance liquid chromatographic
system equipped with a UV detector set at 220 nm and an ODS Hypersil column
is used. The method met precision, specificity, and accuracy requirements. The
hormonal levels determined in one target mare throughout pregnancy were 97.91
to 449.13, 116.47 to 266.02, 74.92 to 235.54, and 84.26 to 300.03 ng/mL, reaching
a maximum towards the 25th, 20th, 33rd, and 27th weeks, respectively, for E1,
EL, EQ, and 17dEQ. The method was successfully tested by quantitating these
estrogens in the plasma from a pregnant mare. Its applicability to the study
of estrogen bioavailability and bioequivalence is suggested.
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