Biomedical Chemistry: Research and Methods 2021, 4(2), e00147

Measurement of Breast Tissue Estrogens by Liquid Chromatography-Tandem Mass Spectrometry

D.A. Olisov1, V.V Chagovets2, N. L. Starodubtseva2, A.A. Smetnik2, V.V. Rodionov2, V.V. Kometova2,
K.S. Chingin3, V.E. Frankevich2*

1Skolkovo Institute of Science and Technology, Skolkovo, Russian Federation
2Academician V.I. Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology,
4 bld. 2 Oparina str.,  Moscow, 117513 Russia; e-mail: vfrankevich@gmail.com
3East China University of Technology, Nanchang, China

Keywords:estrogen; breast cance; breast tissue; LC-MS/MS

DOI:10.18097/BMCRM00147

The whole version of this paper is available in Russian.

Although estrogen contribution estrogen to breast cancer development is not fully understood, an effective method of their measurement, in the mammary gland might provide additional insight. In this study, we have developed a LC-MS/MS method of simultaneous quantification of estrone and estradiol in breast tissue samples. Analytes were extracted with methyl tert-butyl ether by sonication and derivatized with dansyl chloride. Estrogens were analyzed by liquid chromatography-tandem mass spectrometry with an electrospray ionization source. Accuracy and precision were better than 20% for most concentrations. Although estrone and estradiol levels in normal and malignant breast tissue samples analyzed using our method insignificantly differed. The method developed may be used in further studies aimed at evaluating a role estrogens in breast cancer risk.

Figure 1. Absolute peak areas of estradiol (A) and estrone (B) in three sections of a single biopsy sample, each using three chopping options before the extraction step. 1 - whole piece, 2 - chopped into large pieces, 3 - finely chopped.
Figure 2. Graduation curves of tested estrogens (A - estrone, B - estradiol). San - peak area of analyte, SIS - peak area of internal standard.
Figure 3. The histogram of normalised peak areas of estrone and estradiol in three sections of breast tissue sample; A - normalised to tissue mass, B - normalised to lipid mass fraction. E1 - estrone, E2 – estradiol.
Figure 4. The distribution of estrogen content in healthy tissue and neoplasia samples.

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Table 1. Mass spectrometric analysis conditions for each analyte.

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Table 2. Normalized estrogen peak areas for each section and chopping type. Using Student's t-test the comparison was made for each type of chopping.

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Table 3. Reproducibility and accuracy of the methodology. RSD - relative standard deviation, RME - mean error of approximation.

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Table 4. Normalised peak areas of estrone and estradiol analysed in 3 sections of a single biopsy.

FUNDING

This work was supported by the RFBR and the National Natural Science Foundation of China under Project No. 19-515-55021.

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