Soy isoflavone metabolism in cats compared with other species: urinary metabolite concentrations and glucuronidation by liver microsomes

Abstract

1. Soybean is a common source of protein in many pet foods. Slow glucuronidation of soy-derived isoflavones in cats has been hypothesized to result in accumulation with adverse health consequences. Here, we evaluated species' differences in soy isoflavone glucuronidation using urine samples from cats and dogs fed a soy-based diet and liver microsomes from cats compared with microsomes from 12 other species. 2. Significant concentrations of conjugated (but not unconjugated) genistein, daidzein and glycitein, and the gut microbiome metabolites, dihydrogenistein and dihydrodaidzein, were found in cat and dog urine samples. Substantial amounts of conjugated equol were also found in cat urine but not in dog urine. 3. β-Glucuronidase treatment showed that all these compounds were significantly glucuronidated in dog urine while only daidzein (11%) and glycitein (37%) showed any glucuronidation in cat urine suggesting that alternate metabolic pathways including sulfation predominate in cats. 4. Glucuronidation rates of genistein, daidzein and equol by cat livers were consistently ranked within the lowest 3 out of 13 species' livers evaluated. Ferret and mongoose livers were also ranked in the lowest four species. 5. Our results demonstrate that glucuronidation is a minor pathway for soy isoflavone metabolism in cats compared with most other species.

Keywords: Cat; daidzein; dog; equol; genistein; glucuronidation; isoflavone; soy.

Figure 1

Chemical structures of genistein, dihydrogenistein, daidzein, dihydrodaidzein, glycitein, and equol. Also shown for reference are the positions of hydroxyl groups (7-hydroxyl and 4’-hydroxyl) that may be glucuronidated.

Figure 2

Comparison of the rates of glucuronidation of genistein (A: 7-OH-glucuronidation, B: 4’-OH-glucuronidation), daidzein (C: 7-OH-glucuronidation, D: 4’-OH-glucuronidation), equol (E: 7-OH-glucuronidation), and estradiol (F: 3-OH-glucuronidation) by liver microsomes prepared from 13 different species. Species are ranked from lowest (left) to highest (right) for each glucuronidation activity. Glucuronidation of the endogenous substrate, estradiol, was included as a positive control. In all species, 7-OH-glucuronidation of genistein and daidzein predominated over 4’-OH-glucuronidation, while equol 4’-OH-glucuronidation was not detected. Cat liver microsomes (indicated by solid arrow) consistently ranked in the lowest 3 species for 7-OH-glucuronidation of all the isoflavones, while they were the middle ranked species in estradiol glucuronidation. Other species that also consistently ranked low in isoflavone 7-OH-glucuronidation included ferret and mongoose (indicated by dashed arrows). Each bar represents the mean and error bars are the standard deviation of triplicate measurements of pooled liver microsomal samples. The numbers of individual animals contributing to each microsomal pool are indicated on the x-axis of each plot.

Figure 3

Enzyme kinetic plots showing the effect of increasing substrate concentration on 7-OH-glucuronide formation rates measured by HPLC from genistein (A–C), daidzein (D–F), and equol (G–I) for pooled liver microsomes from five cats (A,D,G), 16 dogs (B,E,H), and 48 humans (C,F,I).

Figure 4

Variability in glucuronide formation rates between liver microsomes prepared from 16 different cats. Each filled circle represents the average of duplicate determinations of glucuronide formation rates (7-OH-glucuronidation for isoflavones and 3-OH-glucuronidation for estradiol) for an individual cat measured using 35 µM genistein, daidzein, or equol, or 100 µM estradiol substrate concentration.