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Urinary steroid profiles in pregnancy by capillary column gas chromatography.

A qualitative and quantitative method for the estimation of urinary steroids during pregnancy by capillary column gas chromatography has been developed. Steroid conjugates were hydrolysed by enzymes and free steroids isolated by extraction using SEP-PAK C18 cartridges. Methoxime-trimethylsilyl derivatives of the steroids were characterized by packed column gas chromatography-mass spectrometry and subsequently by their retention indices on capillary column gas chromatography. Of the 48 peaks resolved on the profiles about 35 were identified as steroids. The retention indices of reference steroids were very reproducible in the short term showing a mean coefficient of variation of 0.015%. The columns allowed 95% valley resolution of steroids whose retention indices differed by only 7 units. The use of an on-column injection technique contributed to the high precision for replicate injections (coefficient of variation less than 1%) while for reference steroids, and well-resolved peaks in the urine profile, the mean coefficient of variation for the complete assay was less than 10%. To the best of our knowledge this is the first report concerning the quantitation, with good precision, of a large number of steroids in pregnancy urine.

Chromatography, Gas↗

Steroid profiles during spawning in male common carp.

The profiles of testosterone (T) 11-ketotestosterone (11-KT), 17 alpha-hydroxy-4-pregnen-3-one (17-P), and 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17,20 beta-P) were determined in groups of spermiating male common carp (Cyprinus carpio) which were or were not mated with females and subjected to an increase in water temperature, a treatment which usually induces females to spawn. The steroid levels were correlated with changes in gonadotropin (GtH). All males mated with ovulatory females showed a GtH surge and spawned. Two distinct classes of these spawned males were differentiated based on their initial androgen levels and hormone profiles. Androgen levels were initially elevated in the "high" males. In these fish there was a very rapid shift in the steroidogenic pathway from androgen to progestogen production coincident with the GtH surge. In the "low" males, 17-P levels rose significantly in response to the GtH surge, but there was no steroidogenic shift. In the males mated with nonovulatory females, the levels of 17-P, 17,20 beta-P, T, and 11-KT all increased coincident with a small rise in GtH; there was no steroidogenic shift. In the unmated males, the steroid profiles were similar to those in the nonspawned fish, except that T and 11-KT levels were significantly lower in comparison. In all groups, 17-P rose higher than 17,20 beta-P, and 11-KT was generally the predominant androgen. A model is proposed which explains the steroidogenic shift in the "high" males. This involves the inhibition of androgen biosynthesis by 17,20 beta-P or by a related progestogen.

Androgens↗

Inhibin and relaxin concentrations in early singleton, multiple, and failing pregnancy: relationship to gonadotropin and steroid profiles.

OBJECTIVE: To examine the inter-relationships between inhibin, relaxin, steroid concentrations, estradiol (E2), progesterone (P), and gonadotropins in early pregnancy. DESIGN: Hormone concentrations in plasma were measured during the luteal phase of subjects who became pregnant (n = 58) or failed to become pregnant (n = 47) after ovarian hyperstimulation for in vitro fertilization-gamete intrafallopian transfer (IVF-GIFT) (group 1). A further group of subjects became pregnant (n = 7) or failed to become pregnant (n = 8) during endocrinology tracking of a natural cycle (group 2). Blood was obtained every 3 days in the luteal phase from day 5 in group I (day 0 was oocyte recovery) and from day 0 (first increase in luteinizing hormone [LH]) in group II. RESULTS: Progesterone and E2 were increased over nonpregnant values by day 11 (P) and day 16 (E2) in group I and by day 11 (E2 and P) in group II. Inhibin and relaxin concentrations were significantly increased by day 16 in group I (often by day 11) and by day 14 in group II pregnancy subjects. A direct relationship existed between inhibin, P, relaxin, and human chorionic gonadotropin (hCG). Subjects who had twin pregnancies demonstrated higher concentrations of all hormones and often exhibited increases earlier (by day 11 in group I) than singleton pregnancy subjects. Pregnancies that ended in miscarriages tended to have lower concentrations of P and inhibin. None of the hormones reliably discriminated between the clinical conditions of blighted ovum and of spontaneous abortion, and the predictive value of any hormone measured for miscarriage was not high. CONCLUSIONS: The trend of inhibin and relaxin concentrations closely parallels rises in P during early pregnancy. Luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels are suppressed very early in pregnancy. The suppression of LH and FSH in hyperstimulated cycles is more governed by E2 than inhibin in stimulated cycles. Some subjects destined to miscarry exhibit abnormal endocrine changes very early in the luteal phase.

Abortion, Spontaneous↗

Steroid profiles in hydatidiform mole associated with theca-lutein cysts.

Neutral steroids in urine were determined quantitatively with gaschromatography on capillary columns in a case of benign hydatidiform mole associated with bilateral theca-lutein cysts. A remarkable finding was the very high levels of 17-hydroxypregnanolone and pregnanetriol, which continued to rise until the 15th day after molar evacuation.

Adult↗

Hormonal status of breast cancer. I. Thoretical basis for the analysis of steroid profile.

Gas-liquid chromatography was applied to the analysis of urinary steroids of women. In the statistical analysis, quantitation of all steroid excretions and ratios among them were transformed to give a normal frequency distribution. The influence of the host's age and the menstrual cycle on 24 hours' excretion was investigated for each of 14 identified neutral steroids, of which four in our fraction 2 (11-hydroxyandrosterone, 11-hydroxyetiocholanolone, pregnanediol, and pregnanetriol) were menstruation dependent. The 14 neutral steroid excretions were generally correlated with one another within the host, and the use of a steroid ratio rather than quantitation of steroid excretion was often more sensitive in the characterization of the hormonal status.

11-Hydroxycorticosteroids↗

Effects of ethanol on steroid profiles in the rat testis.

The effects of ethanol on the concentrations of steroids in testis was studied in adult rats. Testosterone, seven of its potential precursors, three of its metabolites, and estradiol were analyzed by gas chromatography-mass spectrometry of samples from testes removed 2 h after intraperitoneal administration of ethanol, 1.2 g/kg body weight. The same analyses were made on samples from control rats. Ethanol gave a marked increase of all 3 beta-hydroxy-delta 5 steroids analyzed: pregnenolone (60%), 17-hydroxypregnenolone (480%), dehydroepiandrosterone (430%) and 5-androstene-3 beta, 17 beta-diol (60%). This resulted in highly significant increases of the 3 beta-hydroxy-delta 5/3-oxo-delta 4 steroid ratios for all steroid couples analyzed. An analogous increase of the ratio between 5 alpha-androstane-3 beta, 17 beta-diol and dihydrotestosterone was also observed, whereas the ratio between androstenediol and dehydroepiandrosterone was decreased by ethanol. The concentration of estradiol was not affected. The results indicate that moderate doses of ethanol inhibit the conversion of 3 beta-hydroxy-delta 5 to 3-oxo-delta 4 steroids. This may be one mechanism by which ethanol decreases the production of testosterone.

Androgens↗

Automated urinary steroid profiles by capillary column gas-liquid chromatography and a computing integrator.

13 Urinary steroid metabolites have been determined by automated gas-liquid chromatography with a 20-m glass capillary column and a computing integrator. Concentrations up to 2 mg/24 h computed by the integrator compare well with those obtained by peak height measurements. At higher concentrations discrepancies occurred, paticularly for the C21 steroids where falsely low values were calculated using peak heights. Mean excretion by healthy males and females of seven steroid metabolities is presented.

Aged↗

[A case of virilizing adrenal cortical adenoma whose steroid profile evokes a partial 21-hydroxylase deficiency (author's transl)].

The C19 and C21 urinary steroids from a virilizing adrenal tumour with high levels of plasma 17 alpha-hydroxyprogesterone and its urinary metabolites have been identified and quantitated gas chromatography and mass spectrometry of sephadex fractions of the total urinary extract. A of the fifty-five identified steroids thirteen were compounds not found before in such a case. The actiology of the apparent 21-steroid hydroxtlase deficiency is discussed at the light of these analytical results and of the hormonogenesis enzymatic induction of the tumour biopsy.

Adenoma↗

Steroid profiles in cultured female jundiá, the Siluridae Rhamdia quelen (Quoy and Gaimard, Pisces Teleostei), during the first reproductive cycle.

The jundiá Rhamdia quelen (Quoy and Gaimard) is a teleost species from the Siluridae family and is an important species for aquaculture in temperate and subtropical climates. Gonad and blood tissue samples were taken from cultured jundiá females between 1998 and 1999. Plasma concentrations of 17beta-estradiol (E(2)), testosterone (T), 11-ketotestosterone (11-KT), 17-hydroxy-4-pregnene-3,20-dione (17-P), 17,20beta-dihydroxy-4-pregnen-3-one (17,20beta-P), and 17,20beta,21-trihydroxy-4-pregnen-3-one (20beta-S) were measured by radioimmunoassay and potential correlations with the stage of oogenesis and sexual maturation examined. During the experimental period two spawning episodes were observed. Plasma concentrations of E(2) increased progressively during oocyte development, simultaneously with the appearance of yolk vesicles and increasing amounts of deposited yolk. In female jundiá, the T peak occurred in October and was coincident with the peak in gonadosomatic index. Two distinct peaks of progestogens were detected, corresponding to the two spawning episodes, suggesting that one or more of these steroids might act as the "maturational-inducing steroid" in jundiá. Unusually large amounts of 11-KT were also measured in the plasma of mature jundiá females. The identity of 11-KT was confirmed by thin-layer chromatography. Although the profiles of the other steroids are compatible with the roles proposed for the action of these hormones in other teleosts, the role of 11-KT, normally found only in males, is unknown.

Animals↗

The plasma sex steroid profiles in the freshwater, sex-reversing teleost fish, Monopterus albus (Zuiew).

The plasma levels of both the free and conjugated forms of six sex steroids (androstenedione, testosterone, 11-oxotestosterone, 11 beta-hydroxytestosterone, estrone, and 17 beta-estradiol) were determined by radioimmunoassay combined with Celite chromatography in different sexual phases of the protogynous Monopterus albus throughout the reproductive cycle. The amounts of 11-oxotestosterone and 11 beta-hydroxytestosterone were found to be very low and variable in all the specimens investigated. No conjugated 17 beta-estradiol or free estrone was detected. Female individuals showed a prespawning rise of androstenedione which subsequently dropped to a low level in the spawning period. The estrogen levels in the female phase were found to be higher than those in all other sexual phases during the spawning period, but the testosterone level remained constant throughout the reproductive cycle in the female phase. The level of androstenedione was highest in the early intersexual and mid-intersexual phases during the postspawning/inactive period. Compared with the female specimens in the same reproductive period, the early intersexual individuals showed a higher level of 17 beta-estradiol, while the mid-intersexual animals showed a higher level of testosterone in the postspawned/inactive period. In the mid-intersexual phase, the levels of androstenedione, testosterone, 17 beta-estradiol, and estrone dropped progressively in relation to the seasonal reproductive cycle. The male fish had a constant level of androstenedione, estrone, and 17 beta-estradiol. However, the level of testosterone increased as the spawning period approached. The hormonal profile in the late intersexual phase was essentially similar to that in the male phase. The changes in the plasma levels of sex steroids in M. albus apparently were related to the maturation of the female and male sex tissues and to their seasonal reproductive cycle.

Androstenedione↗

Steroid profiles of brown adipose tissue.

In brown adipose tissue of alp-marmot (Marmota marmota), badger (Meles meles) and Wistar rats steroids of C21- and C19-type are identified and quantified. The detection of 3 alpha-hydroxy-5 alpha-pregnan-20-one, 3 alpha-hydroxy-5 beta-pregnan-20-one, 3 alpha,21-dihydroxy-5 alpha-pregnan-20-one, 3 alpha,21-dihydroxy-5 beta-pregnan-20-one and 3 beta,21-dihydroxy-5 alpha-pregnan-20-one is of special interest since sleep-inducing properties have been described with these steroids.

Adipose Tissue, Brown↗

Faecal steroids and colorectal cancer: steroid profiles in subjects with adenomatous polyps of the large bowel.

In this necroscopy study the relation between carriage and size of colorectal polyps was correlated with luminal steroid concentrations in respect to malignant risk. Of the 92 subjects entered into the study, 68 had adenomatous polyps of the large bowel, of which 19 had adenomas > 0.9 cm in diameter (large adenomas), 26 in the range 0.5-0.9 cm in diameter (medium adenomas) and 23 of 0.4 cm or less in diameter (small adenomas). Sixty-three percent of subjects carrying large adenomas and 26% of persons carrying small adenomas had an abnormal ratio (> 1.0) of lithocholic acid to deoxycholic acid in intestinal contents as compared to 17% of the adenoma-free comparison group (n = 24). These findings support the suggestion that the ratio of lithocholic acid to deoxycholic acid as a faecal marker may be a useful adjunct to screening procedures for colorectal cancer.

Adenoma↗

Blood steroid profile and in vitro steroidogenesis by ovarian follicles and testis fragments of adult sea lamprey, Petromyzon marinus.

The main purpose of the study was to identify the principal gonadal steroids synthesized by male and female sea lampreys, Petromyzon marinus. To achieve this, we used high performance liquid chromatography to separate the steroids in the serum of sexually mature animals, and to separate the steroids produced by gonadal tissue incubated in the presence of radiolabelled precursor steroids, as a means of identifying the major steroidogenic pathways. We were unable to detect evidence of the 'classical' steroids, such as 17beta-estradiol (E(2)) or testosterone (T) in the serum of either male or female lampreys. Instead, the principal chromatographic peaks contained very polar compounds that had elution times consistent with 15alpha-hydroxylated estrogens and androgens, and there were sex-specific differences in the chemical nature and the quantity of these compounds. Testis fragments or ovarian follicles co-incubated with tritium-labelled pregnenolone ([3H]P(5)), 17-hydroxyprogesterone ([3H]17OHP(4)), or androstenedione ([3H]A(4)), provided additional confirmation that the gonads synthesize a range of very polar steroids, and the metabolites found were consistent with the presence of a 15alpha-hydroxylated (15alphaOH) metabolic pathway common to testis and ovary. For ovarian tissue, the major 'end product' metabolites from all three precursors were 15alphaOH-estrogens, and for testis tissue 15alpha-hydroxyprogesterone (15alphaOHP(4)) and 15alpha-hydroxytestosterone (15alphaOHT) and small amounts of 15alphaOH estrogen. Small amounts of E(2) were also produced by both ovarian (all substrates) and testicular tissue (some substrates). Although it was assumed that the E(2) was synthesized via the aromatization of T, [3H]T was not found as an intermediate metabolite. The study suggests that the principal gonadal steroids in sea lamprey are 15alpha-OH compounds, and that only small amounts of E(2) or T are synthesized by the gonads at this stage of reproductive development. There was no direct evidence of progesterone (P(4)) synthesis from [3H]P(5), although the metabolites synthesized by both testis and ovary were indicative of a metabolic pathway that involved P(4) as an intermediate.

Androgens↗

The steroid profile of a virilizing ovarian tumor.

A case report of a 25-year-old female with a sex cord stromal virilizing ovarian tumor is presented. The pathway of ovarian steroid secretion in this tumor is elucidated with the dominant elements being pregnenolone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, androstenedione, and testosterone. The tumor primarily made testosterone (T) with lesser elevations of androstenedione (A), dehydroepiandrosterone (DHEA), and dihydrotestosterone (DHT). Expert pathologic opinions differed whether this neoplasm was a Sertoli-Leydig tumor or a virilizing granulosa tumor; therefore, it was probably a gynandroblastoma. A unilateral salpingo-oophorectomy was performed and the patient promptly resumed normal ovarian function with ovulation.

17-alpha-Hydroxypregnenolone↗

[Steroid profiles in patients with amenorrhea during induction of ovulation with HMG].

Serum concentrations of various hormones in seven normal women were measured daily for 5 days before and after ovulation. Steroid levels were also measured in severe amenorrheic patients during the induction of ovulation with HMG-HCG. Blood samples from the patients of II grade amenorrhea were collected on the day when the cervical mucus increased more than 200 mm3 in HMG therapy. HCG was given after the blood samples were obtained. Ovulation was successfully induced in six patients and they were classified as group I. In 8 patients induction of ovulation did not succeed and these patients were classified as group II. Hormone levels including LH, FSH, estradiol (E2), progesterone (P4), 17 alpha OH-P4 (17P4), delta 4 androstenedione (delta 4 A), testosterone (Tes.), pregnenolone (P5), 17 alpha OH-P5 (17P5), DHA, delta 5 androstenediol (delta 5 AD), and 20 alpha OH-P4 (20P4) were measured by specific RIA. The following results were obtained. Steroid levels during normal ovulatory cycle: Levels of E2 (380 +/- 16 pg/ml), P5 (6.9 +/- 4.1 ng/ml), and Tes. (3.3 +/- 1.2 ng/ml) showed a peak on the day before LH surge. A significant increase in P4, 17P5 and 20P4 levels was observed after ovulation. Hormone levels in group I: FSH in group I was significantly higher while LH was lower than that in normal women measured during -1 to -3 days from LH surge. On the other hand, among the steroids measured, significantly low Tes. and high 17P5, and E2 levels were noticed in group I. Comparison of hormone levels between group I and II: FSH and LH levels showed no significant difference between the two groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Amenorrhea↗

Intratesticular and plasma steroid profiles in fertile and infertile men.

In 24 apparently healthy men who requested sterilization by vasectomy, we have measured the intratesticular and peripheral plasma levels of 9 unconjugated and 5 sulphoconjugated steroids. These steroid levels were then compared with those found in a group of 18 men, admitted to the hospital with complaints of infertility. With the exception of higher dihydrotestosterone levels in the infertile group, no differences were found between the two groups with regard to the peripheral plasma levels of the unconjugated or sulphoconjugated steroids studied. On the other hand, the intratesticular levels of cortisol, pregnenolone, androstenedione, dihydrotestosterone, pregnenolone sulphate and testosterone sulphate (but not those of testosterone) were significantly diminished in the infertile group. With the exception of dehydroepiandrosterone levels in the fertile group, and dehydroepiandrosterone sulphate and dihydrotestosterone sulphate in the infertile group, the intratesticular steroid levels were uncorrelated with the plasma levels. The data are interpreted as suggesting that a) male infertility may be associated with intratesticular alterations of steroid metabolism which are not reflected by the peripheral steroid levels; b) the plasma and intratesticular levels of testosterone are not sensitive enough indices for the endocrine evaluation of male infertility; c) the role of the sulphoconjugated forms of steroids in the maintenance of male fertility deserves further investigation.

Adult↗

Incomplete androgen insensitivity: asymmetry in morphology and steroid profile and metabolism of the gonads. An analysis of a case.

A patient with clinical manifestations of the incomplete androgen insensitivity syndrome was studied with respect to peripheral blood levels of steroids and steroid sulphates before, during and after gonadectomy. Steroid and steroid sulphate concentrations were also analyzed in spermatic venous blood and gonadal tissue collected during surgery. The metabolic capacity of gonadal tissue was also studied in vitro using progesterone, dehydroepiandrosterone sulphate and oestrone sulphate as substrates. Profound differences between the two gonads were noted with respect to both steroid content and release into pampiniform veins and to in vitro conversion of progesterone and oestrone sulphate. Histological examination revealed the presence of seminiferous tubules with carcinoma in situ in both gonads. It is suggested that the differences between the gonads may be due to an autonomous steroid production in the right gonad in spite of adequate or even elevated gonadotrophic stimulation resulting in a steroidogenic situation resembling the complete androgen insensitivity syndrome, while the conditions found in the left gonad more resembles the incomplete form of the disease.

Adenoma↗