Carbon Isotope Ratio Analysis of Steroids by High-Temperature Liquid Chromatography-Isotope Ratio Mass Spectrometry

Abstract

Generally, compound-specific isotope analysis of steroids is carried out by gas chromatography combined with isotope ratio mass spectrometry. Thus, a derivatization of the steroids prior to the measurement is compulsory, and a correction of the isotopic data is often necessary. To overcome this limitation, we present a new approach of high-temperature liquid chromatography coupled with photodiode array detection and isotope ratio mass spectrometry (HT-LC/PDA/IRMS) for the carbon isotope ratio analysis of unconjugated steroids. A steroid mixture containing 19-norandrosterone, testosterone, epitestosterone, androsterone, and 5β-pregnane-3α,17α,20α-triol was fully separated on a C4 column under high-temperature elution with water as the sole eluent. The accuracy for isotope analysis (±0.5 ‰) was around 20 μg g(-1) for testosterone, epitestosterone (79 ng steroid absolute on column), and 30 μg g(-1) for 19-norandrosterone, androsterone, and 5β-pregnane-3α,17α,20α-triol (119 ng steroid absolute on column). The applicability of the method was tested by measuring a pharmaceutical gel containing testosterone. With this work, the scope of LC/IRMS applications has been extended to nonpolar compounds.

Determination of carbon isotope ratios for honey samples by means of a liquid chromatography/isotope ratio mass spectrometry system coupled with a post‐column pump

“…Despite these problems, LC/IRMS has been used to analyze compounds in various sample media, including caffeine in beverages; 13 organic acids, glucose, and fructose in lime and lemon juices; 14,15 glycerol and ethanol in wine; [16][17][18] vanillin in chocolate bars and snack foods; 19 glyphosate and aminomethylphosphonic acid; 20 sulfonamides in pharmaceuticals; 21 glucose in blood plasma; 22 amino acids; 23,24 steroids; 25 and sugars (glucose, fructose, disaccharides, trisaccharides, and oligosaccharides) in honey. [26][27][28][29] Honey is a highly nutritious functional food composed mainly of glucose, fructose, minor sugars, organic acids, amino acids, vitamins, and minerals.…”

Annual banned‐substance review: analytical approaches in human sports drug testing

“…Zhang et al . demonstrated the utility of LC/IRMS in separating 19‐norandrosterone, T, EpiT, A, E, and 5β‐pregnane‐3α,17α,20α‐triol using high‐temperature LC and determining respective carbon isotope ratios, which might open new possibilities of sports drug testing in the future …”

“…Zhang et al demonstrated the utility of LC/IRMS in separating 19-norandrosterone, T, EpiT, A, E, and 5β-pregnane-3α,17α,20α-triol using high-temperature LC and determining respective carbon isotope ratios, which might open new possibilities of sports drug testing in the future. [69] Alternative matrices and complementary approaches Detection possibilities concerning AAS in alternative matrices have been assessed with particular attention being paid to testosterone and its esterified derivatives. Since transdermal testosterone applications have been shown to result in only modest alterations of the urinary steroid profile, the option to use oral fluid was tested by Thieme et al [70] Following the transdermal administration of approximately 3-7 mg of testosterone, salivary concentrations of the target analyte increased from baseline (30-142 pg/mg) to more than 1000 pg/mg over a period of 16 h, suggesting a viable means to efficiently screen for the misuse of transdermal testosterone preparations.…”

A novel high‐temperature combustion interface for compound‐specific stable isotope analysis of carbon and nitrogen via high‐performance liquid chromatography/isotope ratio mass spectrometry

“…amino sugars) and sports (e.g. doping; steroids) . A number of these disciplines utilize standard analytical CSIA techniques.…”

“…Possible fractionation must also be controlled in derivatization for subsequent GC/IRMS measurements. Both EA/IRMS and GC/IRMS CSIA also require additional corrections, which increase the uncertainty of the determined values …”