Steroids quantitation

Clearly, there are basic successful practices evolving with regard to MS/MS steroid quantitation, which can be summarized as follows ... 

There are no reports on steroid quantitation by LC-MS, but cortisol, DHEA, and testosterone are very low when measured by RIA, and DOC and corticosterone are elevated. With the exception of DOC, all of these analyte assays are available by MS/ MS, as are progesterone and pregnenolone, which should also be elevated in the condition. 

The unsaponifiable part of edible fats contains a series of cyclic triterpenes which have structures related to that of steroids. Quantitatively, the 3 P-hydroxysteroids are the sterols which are to the fore. Especially diverse is the sterol spectrum of plant fats which contain not only desmethyl, but also 4-methyl and 4,4-dimethyl sterols. 

GC-MS methods have also been used for steroid quantitation but these usually require extraction and purification steps along with derivatization of the steroid before measurement, which complicates the process and involves time. 

D-Mevalonic acid is the fundamental intermediate in the biosynthesis of the terpenoids and steroids, together classed as poly-isoprenoids. The biogenetic isoprene unit is isopentenyl pyrophosphate which arises by enzymic decarboxylation-dehydration of mevalonic acid pyrophosphate. D-Mevalonic acid is almost quantitatively incorporated into cholesterol synthesized by rat liver homogenates. 

Two approaches to convergent steroid syntheses are based on the thermal opening of benzocyclobutenes to the o-quinodimethane derivatives (see p. 80 W. Oppolzer, 1978 A) and their stereoselective intramolecular Diels-Alder cyclizations. T, Kametani (1977 B, 1978) obtained (+ )-estradiol in a six-step synthesis. The final Diels-Alder reaction occurred regio- and stereoselectively in almost quantitative yield, presumably because the exo transition state given below is highly favored over the endo state in which rings A and D would stcrically inter-... 

This publication provides several examples of the use of solid-phase extractions for separating analytes from their matrices. Some of the examples included are caffeine from coffee, polyaromatic hydrocarbons from water, parabens from cosmetics, chlorinated pesticides from water, and steroids from hydrocortisone creams. Extracted analytes maybe determined quantitatively by gas (GC) or liquid chromatography (LG). 

Quantitative Structure—Activity Relationships. Many quantitative stmcture—activity relationship (QSAR) studies of progestins have appeared in the Hterature and an extensive review of this work is available (174). QSAR studies attempt to correlate electronic, steric, and/or hydrophobic properties to progestational activity or receptor binding affinity. A review focusing on the problems associated with QSAR of steroids has been pubUshed (175). 

S. Goxo, Quantitative Analysis of Steroids, Elsevier Scientific Publishing Co., Amsterdam, the Netherlands, and Akademiai Kiado, Budapest, Hungary, 1983. 

The kinetics of formation and hydrolysis of /-C H OCl have been investigated (262). The chemistry of alkyl hypochlorites, /-C H OCl in particular, has been extensively explored (247). /-Butyl hypochlorite reacts with a variety of olefins via a photoinduced radical chain process to give good yields of aUyflc chlorides (263). Steroid alcohols can be oxidized and chlorinated with /-C H OCl to give good yields of ketosteroids and chlorosteroids (264) (see Steroids). /-Butyl hypochlorite is a more satisfactory reagent than HOCl for /V-chlorination of amines (265). Sulfides are oxidized in excellent yields to sulfoxides without concomitant formation of sulfones (266). 2-Amino-1, 4-quinones are rapidly chlorinated at room temperature chlorination occurs specifically at the position adjacent to the amino group (267). Anhydropenicillin is converted almost quantitatively to its 6-methoxy derivative by /-C H OCl in methanol (268). Reaction of unsaturated hydroperoxides with /-C H OCl provides monocyclic and bicycHc chloroalkyl 1,2-dioxolanes. 

The QSAR (quantitative structure-activity relationship) approach has been considered for the identification of toxicants that bind to steroid and aryl... 

The same qualitative results based on steric and electronic properties of the steroid ring system are found regardless of the structure of the peracid. Thus, although the generality of stereochemistry can be discussed with regard to all peracids, a quantitative comparison is valuable in some instances. 

The properties of chlorine azide resemble those of bromine azide. Pon-sold has taken advantage of the stronger carbon-chlorine bond, i.e., the resistance to elimination, in the chloro azide adducts and thus synthesized several steroidal aziridines. 5a-Chloro-6 -azidocholestan-3 -ol (101) can be converted into 5, 6 -iminocholestan-3l -ol (102) in almost quantitative yield with lithium aluminum hydride. It is noteworthy that this aziridine cannot be synthesized by the more general mesyloxyazide route. Addition of chlorine azide to testosterone followed by acetylation gives both a cis- and a trans-2iddMct from which 4/S-chloro-17/S-hydroxy-5a-azidoandrostan-3-one acetate (104) is obtained by fractional crystallization. In this case, sodium borohydride is used for the stereoselective reduction of the 3-ketone... 

The reaLUons of phenyltetrafluorophosphorane with numerous silylated secondary or tertiary a- or (1-hydroxy esters, ketones, nitriles, ethers, nitro, and trichloromethyl derivatives have been investigated, the corresponding a or p fluoro derivatives are obtained in yields varying from reasonable to nearly quantitative [24, 25, 26, 27 The application of phenyltetrafluorophosphorane for fliiorination of silyloxy steroids has also been reported [28]... 

Pentafluorobenzyl bromide has been used in the derivatization of mercaptans [55] and phenols [36], m the analysis of prostaglandins [37], and in quantitative GC-MS [5S] 1,3 Dichlorotetrafluoroacetone is used for the derivatization of amino acids to the corresponding cyclic oxazolidinones and allows the rapid analysis of all 20 protein ammo acids [d] Pentafluorophenyldialkylchlorosilane derivatives have facilitated the gas chromatographic analysis of a wide range of functionally substituted organic compounds, including steroids, alcohols, phenols, amines, carboxylic acids, and chlorohydrms [4]... 

The [4-1-3] cycloaddition has also been realized in acceptors containing a nitrogen atom. While a,/ -unsaturated aldimines, and structurally flexible ketimine such as (87), generally only undergo [3-1-2] cycloadditions (see Scheme 24), the ketimine (112), which is rigidly held in a cisoid conformation, does give exclusively the [4-1-3] adduct azepine (113). On the other hand, the steroidal imine (114) produces a quantitative yield of a 1 1 mixture of the [4-1-3] and [3-1-2] cycloadducts (115) and (116), respectively (Scheme 2.31) [36]. 

Note The derivatized steroids can be extracted from the blue chromatogram zones with alcohol and quantitatively determined by means of the Zimmermann reaction, which is not interfered with by the presence of phosphoric acid and phosphomolybdic acid. A yellow background can be bleached by exposure to anunonia vapor [2]. 

A quantitative description of human skin permeabiUty (kp) based on H-bond donor and acceptor factors was obtained with 22 alcohols and steroids [63] ... 

Electron impact ionization, also known as particle beam ionization, has been applied to the online determination of steroids such as hydrocortisone, cortisone, prednisolone and prednisone. Polymer additives such as NC-4, Irga-nox 1076,1-octadecanol and Naugard -XL were identified and quantitated online by electron impact and, separately, by atmospheric pressure chemical ionization methods.78... 

The perturbation of monolayers with agents (e.g., disodium ethylenediamine tetraacetate, Ca+2-free medium, sodium citrate, cytochalasin D) to open tight junctions and the effect on the transmonolayer flux of permeants are addressed in this section. It has been observed that permeants taking predominantly the trans-cellular route are not affected by perturbants of the paracellular route, compared to extracellular or relatively hydrophilic permeants (Artursson and Magnusson, 1990). Let us put these general observations into a quantitative intepretation in the light of the transmonolayer kinetic studies of steroids in this section and of paracellular permeants in Section III. There are three cases to consider (1) ABL-controlled permeants, (2) monolayer-controlled permeants transported principally by the transcellular route, and (3) monolayer-controlled permeants for which the paracellular route dominates. 

Komiya I, JY Park, NFH Ho, WI Higuchi. (1980). Quantitative mechanistic studies in simultaneous fluid flow and intestinal absorption using steroids as model solutes. Int J Pharm 4 249-262. 

A large number of nonenzymatic compounds, including tocopherols, caroti-noids, vitamins C and D, steroids, ubiquinones, thiols, uric acid, bilirubin, ino-sine, taurine, pyruvate, CRP, and so on, demonstrate qualitative antioxidant properties under experimental conditions. However, the quantitative relevance of most findings remains unclear. 

Ferric sulphate is a mild oxidant and is non-reactive with the steroid substrate. It liberates iodine quantitatively (equation 76), and the iodine is extracted into CH2CI2 and consumed as in equation 75. 

Halcinonide has been quantitated in various formulations or as bulk powder by a differential ultraviolet, borohydride reduction assay. 2 This differential assay involves measuring the ultraviolet absorbance of an aliquot of methanolic steroid solution containing sodium borohydride decomposed prior to the addition of steroid. Its absorbance is determined against a methanolic reference solution of steroid reduced by sodium borohydride to destroy the 3-one-4-ene chromophore. The utility of this procedure is that many interferences from excipients and other, unconjugated, steroids can be eliminated in the assay of a formulation. 

A diatomaceous earth column was used to separate halcinonide from excipients . The formulation is mixed with column packing material and then is transferred to the top of the column. Steroidal material is eluted with benzene (CAUTION) under a well-ventilated hood and then quantitated using thin layer chromatography60 0r tetra-methylammonium hydroxide l colorimetry. 

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