Urine 21-hydroxy steroids

Shackleton CH, Irias J, McDonald C, Imperato-McGinley J (1986) Late-onset 21-hydroxy-lase deficiency reliable diagnosis by steroid analysis of random urine collections. Steroids 48 239-250... 

For example, glucocorticoids stimulate gluconeogenesis, lipolysis, and an increased uptake of amino acids by the liver.) Cortisol also possesses a small amount of mineralocorticoid activity. (Mineralocorticoids regulate Na+ and K+ metabolism. For example, aldosterone, the most important mineralocorticoid in humans, induces the reabsorption of Na+ from urine. It also promotes the secretion of K+ and H+ into urine.) For steroids to have either glucocorticoid or mineralocorticoid activity, they must possess a hydroxy group at C-ll. 

One anabohc steroid, the presence of which has proven difficult to analyze, is stanozolol [10418-03-8] C2 H22N20. A metaboUte of the parent dmg, hydroxy stanozolol, detected in equine urine eight hours after ingestion of the parent dmg is actually identified, usually at very low levels. Analysis was done by Ic/ms/ms which had a shortened analysis time advantage over gc/ms procedures because of the elimination of the need for a derivatization step (33). 

Endogenous and exogenous androgens can be derivatized with trimethylsilyl (TMS) for hydroxy functions and by 0-methylation for ketones, and analyzed with GC-FID or GC-MS (Shimada et al., 2001). MS is more prevalent due to unequivocal identification and greatly increased sensitivity but FID is still used in laboratories for some steroids. Sterols have typically been analyzed by GC-FID and GC-MS with derivatization to optimize peak shape (Shimada et al., 2001), and bile acids can be derivatized with M-butyl ester-TMS ether and analyzed by GC-FID from plasma samples (Batta et al., 1998). Juricskay and Telegdy (2000) reported an assay capable of analyzing 28 steroids in urine samples using GC-FID. 

There are three other recognised varieties of disorders leading to congenital adrenal hyperplasia, including 3/3-hydroxysteroid dehydrogenase deficiency. Patients with this defect have grossly elevated levels of 3/3-hydroxy-5-ene steroids in their plasma and urine. Complete lack of the enzyme is incompatible with life however, an incomplete deficiency has been reported [262,263] where GC-MS identification of saturated C19 and C21 steroids revealed that some enzyme activity was present in the liver. 

Leinonen et al. (2002) compared LC-MS-MS with electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI) and atmospheric pressure photoionization (APPI) for unconjugated (free) anabolic steroids in human urinary extracts. The selected analytes were synthetic steroids or their metabolites, known to be misused in sports and known to be excreted in urine unconjugated, namely oxandrolone (Figure 2.1), the 3 -hydroxy metabolite of stanozolol and the 6p-hydroxy metabolite of 4-chlorodehydro-methyltestosterone. 

The important 3y8-hydroxy-A steroids in umbilical cord blood and infant urine are also mainly present as sulfates (E2, E4, Ml, R6, S20) only very small portions of DHA and 16 -OH-DHA, for instance, have been found free in blood (S21). Because of the importance of sulfate conjugation, special care must be taken, if enzyme hydrolysis is used alone, to ensure the presence of the specific sulfatases required. For instance, at least one steroid present in infant urine is diconjugated and cannot be completely hydrolyzed by the enzymes in the crop fluid of... 

The action of the sulfatases in the Helix pomatia enzyme is insufficient to achieve complete hydrolysis at least of the 3/8-hydroxy-A steroids in umbilical cord blood and infant urine, and a further stage involving a solvolytic procedure (B40) is essential (S9, Sll). 

A situation similar to that for urine applies for the assay of the 3 -hydroxy-A° steroids in umbilical cord blood. The amount of blood... 

The adrenal gland is relatively 10-20 times larger than that in the adult and is mainly composed of the fetal zone. After birth involution commences and by day 28 of life the size has decreased by between four-fifths and nine-tenths (P13, Tl). It would not be expected that such a large amount of highly productive tissue would cease activity immediately after birth, and in fact production of steroids of the pattern found in the fetus does not cease until after the sixth month of life (R3, R6, Fig. 7). The concentration of the 3/3-hydroxy-A steroids in urine (Fig. 7) and blood (E4) and of the 17-OS in blood (E4), appears roughly to parallel the involution of the fetal zone of the adrenal (E4). 

Fig. 6. The content of the major 3j8-hydroxy-A steroids in adult male urine, infant urine, term amniotic fluid, and plasma obtained from the umbilical cord at birth.

The 3j8-hydroxy-A steroid content of amniotic fluid (S12) is compared with that in adult and infant urine and umbilical cord blood in Fig. 6. Schindler and co-workers have assayed several 3/3-hydroxy-A steroids and others in amniotic fluid for both normal and abnormal pregnancies (S4, S5). 

The level of DHA in cord blood is very high and the lack of significant quantities of the compound in infant urine may be explained by 16a-hydroxylation being the dominant metabolic pathway (see Section 8 for the excretion of 16a-hydroxy-A steroids). In adults DHA is the principal precursor of androsterone and etiocholanolone, and they are its main metabolites (R11,V1). 

Fig. 7. The urinary excretion of the major 3i8-hydroxy-A steroids and one unknown compound during the first 6 months after birth. Assays were performed on urines from different individuals, each infant being investigated once only.

At least three, 3j8-hydroxy-A steroids have been shown to be present in urine in diconjugated form of these 16 S-OH-DHA and 21-OH-preg-nenolone are hydrolyzed by the Helix pomatia enzyme, but androstenediol (17a) is freed only by solvolysis (Sll, Fig. 5). The proportions of the individual 3j8-hydroxy-A steroids found in umbilical cord plasma mono-and diconjugated are shown in Table 15. Very little is free, but Simmer and co-workers (S21) have established a concentration of free 16a-OH-DHA of approximately 0.4 jag/100 ml. 

Shackleton, C. H. L., and Mitchell, F. L., The measurement of 3/3-hydroxy-A steroids in human fetal blood, amniotic fluid, infant urine and adult urine. Steroids 10, 359-385 (1967). [Note publisher s errors, corrections given in Steroids 11, 415... 

Hydrocortisone (24) (Ri, R, R Xj, X, = H, no A Fig. 15.10) undergoes a variety of oxidative and reductive metabolic conversions (115). Oxidation of its dihydroxyacetone side-chain leads to formation of cortienic acid (25) through a 21-aldehyde (21-dehydrocortisol) and a 21 -acid (cortisolicacid). Cortienic acid is an ideal lead for the inactive metabolite approach because it lacks corticosteroid activity and is a major metabolite excreted in human urine. To obtain active compounds,the important pharmacophores found in the 17a and 17P side-chains had to be restored. Suitable isosteric/isoelectronic substitution of the a-hydroxy and /3-carboxy substituents with esters or other types of functions should restore the original corticosteroid activity and also incorporate hydrolytic features to help avoid accumulation of toxic levels. More than 120 of such soft steroids (24) that resulted from modifications of the 17/3-carboxyl function and the 17o -hydroxy function together with other... 

The anabolic steroid B-nortestosterone was rapidly metabolized by the cells, initially resulting in the formation of norandrosten ione, which was further transformed into the glucuronide of 15a-hydroxy-norandrostenedione (60), The same metabolites were subsequently shown to be present in urine samples of pigs treated with B-nortestosterone. 

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