Placental transfer of the soy isoflavone genistein following dietary and gavage administration to Sprague Dawley rats

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Abstract

Genistein, the principal soy isoflavone, has estrogenic activity and is widely consumed by humans for putative beneficial health effects. The goal of the present study was to measure placental transfer of genistein in rats as a possible route of developmental exposure. Pregnant Sprague-Dawley rats were administered genistein orally, either by diet or by gavage. Concentrations of genistein aglycone and conjugates were measured in maternal and offspring serum and brain using HPLC with isotope dilution electrospray tandem mass spectrometry. Although fetal or neonatal serum concentrations of total genistein were approximately 20-fold lower than maternal serum concentrations, the biologically active genistein aglycone concentration was only 5-fold lower. Fetal brain contained predominately genistein aglycone at levels similar to those in the maternal brain. These studies show that genistein aglycone crosses the rat placenta and can reach fetal brain from maternal serum genistein levels that are relevant to those observed in humans.

Introduction

Genistein, the principal soy isoflavone, has been the subject of numerous studies in experimental animals and humans because of possible beneficial [1] and adverse effects [2], [3] due to estrogenic activity. Genistein has been shown to prevent [4], [5] or to promote [6], [7] chemically induced mammary tumor formation in rodents, depending on the timing of exposure to genistein (prepubertal vs. in utero, respectively) and timing of chemical carcinogen exposure (either dimethylbenzanthracene or methylnitrosourea). Genistein and other isoflavones, primarily as the glucuronide conjugates, have been measured in umbilical cord plasma and amniotic from human pregnancies and the concentrations were comparable to those present in maternal plasma [8]. These reports suggested that human and experimental animal fetuses can be exposed to genistein through maternal consumption. However, Lamartiniere et al. concluded that prenatal exposure of rats to genistein through the maternal diet did not confer protection from chemically induced mammary cancer later in life because conjugated forms of genistein, the principal form in maternal blood, could not cross the placenta [9].

Estrogenic effects of genistein in the central nervous system have been reported in vitro and in vivo: an estrogen-dependent cell line derived from the anterior pituitary showed enhanced release of prolactin [10]; injection in neonatal rats altered neuroendocrine secretions (gonadotropin releasing hormone, luteinizing hormone) and produced developmental changes in the sexually dimorphic nucleus of the hypothalamus [11; Scallet, unpublished); and dietary administration to ovariectomized Sprague-Dawley rats increased prolactin secretion [12]. These findings are also consistent with the hypothesis that genistein can penetrate both the placental and blood brain barriers to a level sufficient to exert estrogenic effects. The exposure of the fetus to genistein is significant because the developing brain may be more susceptible than adult brain to chemically mediated events because of metabolic differences, incomplete formation of the blood-brain barrier, or the irreversibility of developmental effects [13], [14].

Consumption by animals and humans of soy isoflavones, which occur in the plant as glucosides, results in cleavage by microbial β-glucosidases in the gut, absorption of the aglycones, followed by conversion to glucuronide and sulfate conjugates through first pass metabolism in the intestine and liver and release into the circulation [1], [15]. The research described here relates concentrations of genistein, the aglycone and putative hormonally active form, and total genistein, which includes the conjugated forms (primarily glucuronides), in maternal and fetal serum and brain following oral administration of genistein. These determinations provide information during critical early developmental periods about genistein exposure derived from serum concentrations in pregnant female rats that are relevant to human exposures [16].

Section snippets

Reagents

Genistein, with purity greater than 99% determined using 1H- and 13C-NMR, EI/MS, melting point, and TLC analysis, was obtained from Toronto Research Chemicals (Ontario, Canada); crude glucuronidase/sulfatase from Helix pomatia containing 105 units/mL glucuronidase activity + 5 × 103 units/mL sulfatase activity was obtained from Sigma Chemical Co. (St. Louis, MO). Deuterated genistein (6,8,3′,5′-d4, 95%) was purchased from Cambridge Isotope Laboratories (Andover, MA) and characterized previously

Results

Initial studies used neonatal pups analyzed within two days after delivery from pregnant female rats that had been maintained on either soy-free or genistein-fortified feed from birth through adulthood as part of a multigeneration study. The results obtained from analysis of 18 individual pups in eight litters derived from dams consuming 500 μg/g genistein diet and two litters from dams consuming the control diet are shown in Table 1. The method detection limit was approximately 2 nM, which was

Discussion

These studies suggest that genistein crossed the placenta into blood, brain, and probably other organs of the developing rat fetus either by passive uptake of the aglycone or by placental hydrolysis of conjugated forms [20]. This conclusion is at odds with that of Lamartiniere et al. who reported low concentrations of genistein aglycone (ca. 3 nM) in fetal serum following administration through maternal diet [9]. Because LC/MS/MS methodology with similar sensitivity was used to quantify

Acknowledgements

The high level technical support of Connie Weis and Carrie Moland and helpful discussions with Daniel M. Sheehan and William Slikker, Jr., all from NCTR, are gratefully acknowledged. This research was supported in part by Interagency Agreement #224–93-0001 between NCTR/FDA and the National Institute for Environmental Health Sciences/National Toxicology Program. HCC acknowledges support of a fellowship from the Oak Ridge Institute for Science and Education, administered through an interagency

References (25)

  • L. Hilakivi-Clarke et al.

    Prepubertal exposure to zealeranone or genistein reduces mammary tumorigenesis

    Br J Cancer

    (1999)
  • L. Hilakivi-Clarke et al.

    Maternal exposure to genistein during pregnancy increases carcinogen-induced mammary tumorigenesis in female rat offspring

    Oncology Reports

    (1999)
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