Effects of androstenedione administration on epitestosterone metabolism in men

Abstract

Androstenedione is a steroid hormone sold over-the-counter to individuals who expect that it will enhance strength and athletic performance. Endogenous androstenedione is the immediate precursor of testosterone. To evaluate the metabolism of oral androstenedione, we randomly assigned 37 healthy men to receive 0 (group 1), 100 mg (group 2), or 300 mg (group 3) of androstenedione in a single daily dose for 7 days. Eight-hour urines were collected 1 day before the start of androstenedione, and on days 1 and 7. Using gas chromatography-mass spectrometry, we measured excretion rates of glucuronide-conjugated epitestosterone, its putative precursor (E-precursor), and metabolites (EM-1 and EM-2), and we evaluated possible markers of androstenedione administration. Day 1 and 7 rates were not different: the means were averaged. The means (μg/h) for groups 1, 2, and 3, respectively were, for epitestosterone 2.27, 7.74, and 18.0; for E-precursor, 2.9, 2.0, and 1.5; for EM-1/E-precursor 0.31, 1.25, and 2.88; for EM-2/E-precursor 0.14, 0.15, and 1.15; for testosterone/epitestosterone (T/E) 1.1, 3.5, and 3.2. Epitestosterone, EM-1, and EM-2 excretion was greater in groups 2 and 3 versus group 1 (0.0001 < P < 0.03), as were EM-1/E-precursor, EM-2/E-precursor, and T/E. E-precursor excretion was lower in groups 2 (P = 0.08) and 3 (P = 0.047) versus group 1. Androstenedione increases excretion of epitestosterone and its two metabolites, while decreasing that of its precursor. Elevated ratios of EM-1- and EM-2/E-precursor, and the presence of 6α-hydroxyandrostenedione are androstenedione administration markers.

Introduction

Androstenedione, a steroid hormone, is an immediate precursor to testosterone in the intrinsic synthetic pathways of androgens [1], [2]. In the United States, androstenedione is sold over-the-counter and the sport supplement market attained annual sales of 1.4 billion in 1999 [3]. The media suggests that sales continue to grow at a prodigious rate. Androstenedione is marketed primarily to athletes as a potential anabolic agent. Advertising materials claim that androstenedione improves athletic performance, libido, and quality of life; however, none of these assertions have been demonstrated in peer-reviewed studies.

Many sport organizations prohibit the use of androstenedione and some test the urine of athletes for steroids, however the prohibition is difficult to enforce because consensus on the criteria for a positive case of androstenedione use has not been attained. Potential urinary markers of androstenedione administration include extremely high levels of testosterone, DHT, androsterone, and etiocholanolone [4]. In some individuals, androstenedione use increases the ratio of testosterone to epitestosterone (T/E) above the International Olympic Committee (IOC) cut-off of 6 [5], [6], as often occurs in subjects who use testosterone [7]. In those men, the T/E ratio increases because of both an increase in urinary testosterone excretion and a decrease in urinary epitestosterone excretion. Epitestosterone is of great interest in the field of doping control because it is the denominator in T/E, a surrogate marker of testosterone administration. Yet, while there is much information on urinary testosterone, relatively little is known about epitestosterone.

Although epitestosterone was first isolated from urine in 1964 [8], its physiological role has not been established. It has minimal androgenic properties in man [9] and it is produced in both the adrenals and ovaries [8]. The administration of ACTH, hCG, or an i.v. infusion of epitestosterone increases urinary epitestosterone [10]. The production rate of epitestosterone has been estimated at just 200–300 μg/day [10], or 5% of the production rate of testosterone. No precursors of epitestosterone have been conclusively identified, but 5-androsten-3β,17α-diol (E-precursor) has been proposed as a possible precursor [11]. It has also been suggested that epitestosterone might arise as a byproduct of 16-androstene synthesis [12]. In animals, epitestosterone inhibits 5α-reductase and has some anti-androgen activity [13].

We recently demonstrated that oral androstenedione administration increases serum testosterone and testosterone glucuronide levels, and markedly increases the urinary excretion rates of the glucuronides of testosterone, DHT, androsterone, and etiocholanolone [4], [14]. In the same subjects we showed that trace contamination (0.001%) of androstenedione with 19-norandrostenedione is sufficient to cause urine test results positive for 19-norandrosterone, the standard marker for nandrolone use [15]. In this study we examine the effect of oral androstenedione administration on the urinary excretion of glucuronides of epitestosterone, epitestosterone metabolites, and E-precursor. In addition, we examine the effect of oral androstenedione administration on the T/E ratio and discuss diagnostic criteria for the detection of androstenedione administration.