Investigation of urinary steroid metabolites in calf urine after oral and intramuscular administration of DHEA

Abstract

DHEA (3β-hydroxy-androst-5-en-17-one) is a natural steroid prohormone. Despite a lack of information on the effect, DHEA and other prohormones are frequently used as a food supplement by body-builders. DHEA is suspected for growth promoting abuse in cattle as well. Considering the latter, urine samples from a previous exposure study in which calves were exposed to 1 g DHEA per day during 7 days, were used. The calves were divided in three groups: one orally treated, one intramuscularly injected and a control group. The effect of this treatment on the urinary profile of several precursors and metabolites of DHEA was investigated. Urine samples were collected several days before and during the 7 days of administration and were submitted to a clean-up procedure consisting of a separation of the different conjugates (free, glucuronidated and sulphated forms) of each compound on a SAX column (Varian). An LC-MS/MS method was developed for the detection and quantification of several metabolites of the pathway of DHEA including 17α-and 17β-testosterone, 4-androstenedione, 5-androstenediol, pregnenolone and hydroxypregnenolone. Elevated levels of DHEA, 5-androstenediol and 17α-testosterone were observed in the free and sulphated fraction of the urine of the treated calves, thus indicating that the administered DHEA is metabolized mainly by the ∆ 5 -pathway with 5-androstenediol as the intermediate.Sulphoconjugates of DHEA and its metabolites were found to constitute the largest proportion of the urinary metabolites. The free form was also present, but in a lesser extent than the sulphated form, while glucuronides were negligible.

Hair cortisol and dehydroepiandrosterone sulfate concentrations in healthy beef calves from birth to 6 months of age

“…In cattle, it has been reported that cortisol/DHEA ratio increases in lame dairy cows [26] and following transportation of young bulls [28], but decreases in cows with metritis and leucopenia [29], thus revealing conflicting results when evaluating different J o u r n a l P r e -p r o o f stress conditions. Recently, DHEA has been suspected for growth promoting abuse in cattle, and the measurement of DHEA metabolites have been investigated in calf urine [4], in view of the monitoring programs on hormone residues. The assessment of hair DHEA sulfate (HDHEA-S) concentrations in healthy calves may allow not only to investigate possible alteration due to pregnancy disturbances or to poor welfare conditions, but it may represent also a useful tool in the screening strategy to trace abuses with steroids in this species, especially in beef breeds.…”

“…Cortisol is essential for final intrauterine fetal development and maturation [1], as well as to trigger the process of parturition in species like ruminants [2], but also to drive the transition to the extrauterine life [3] which consists in structural and physiological changes from the last intrauterine stage of development until the end of the neonatal period. DHEA is a natural steroid prohormone, and it is a key intermediate in the biosynthesis of biologically potent androgens and estrogens [4]; it is known to exert neuroprotective effects and to play a balancing role against a wide range of C effects [5][6].…”

Urinary Concentrations of Steroids in Bulls under Anabolic Treatment by Revalor-XS® Implant

“…Consequently, several research studies have been implemented through the years to test the effect of different growth promoters administered via different routes [18, 28, 34, 35] and to evaluate residual levels and target metabolites. In this work a commercially available ear implant containing trenbolone acetate and estradiol (Revalor-XS) was tested; this formulation is used in the United States in bovine breeding and is able to release a constant amount of hormones over a time period up to 200 days eliminating the need to reimplant the device.…”

A rapid and sensitive UPLC–MS/MS method for the simultaneous quantification of serum androsterone glucuronide, etiocholanolone glucuronide, and androstan-3α, 17β diol 17-glucuronide in postmenopausal women

“…In most cases, the sample preparation procedures for LC-MS/MS are much simpler than for GC-MS/MS, among which are solid phase, liquid-liquid and protein precipitation extractions [8][9][10]. Therefore, LC-MS/MS appears well-suited for the quantification [11][12][13][14][15][16] and characterization [17][18][19][20][21] of steroid glucuronides in different biological matrices such as urine [8][9][10][11][12][13][14][15][16][17][18][19][20][21], serum [2,3] and tissue [22]. Using liquid-liquid extraction, a highly acidic buffer and polar extraction solvent such as ethyl acetate are required [11], resulting in the co-extraction of many other polar compounds which could potentially interfere with the investigated steroid glucuronides and affect the column life time.…”

“…The previously reported quantitation methods of steroid glucuronides have been applied to the matrix urine at high steroid concentrations [8][9][10][11][12][13][14][15][16][17][18][19][20][21]. In the present work, we develop a fully validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous direct quantitation of androsterone glucuronide (ADT-G), etiocholanolone glucuronide (Etio-G) and androstan-3a, 17b diol 17-glucuronide (3a-diol-17G) in human serum.…”