Endocrine Abstracts (2002) 4 P88

Measurement of urinary 18-hydroxytetrahydro-11-dehydrocorticosterone (18-OHTHA) excretion rate by gas chromatography-mass spectrometry using the heterologous standard, beta-cortol

LA Shakerdi1, JMC Connell1, R Fraser1 & AM Wallace2


1MRC Blood Pressure Group, Western Infirmary, Glasgow; 2Glasgow Royal Infirmary, Glasgow, UK.


Introduction. As for many minor metabolites of steroids, no authentic standard is commercially available for 18-OH-THA. This compound is the principal urinary metabolite of 18-hydroxycorticosterone, a putative intermediate in the synthesis of aldosterone and of diagnostic value in screening for hypermineralocorticoidism. Since de novo synthesis is time-consuming and expensive, we tested the possibility of using beta-cortol, which produces an ion at m/z 457 in common with 18-OH-THA as a standard.

Methods. Urinary steroids were extracted and derivatised as described by Shackleton (1). A GCQ(super)TM Plus benchtop ion trap GC/MS (ThermoQuest, Finnegan, USA) containing a capillary DB-1 column (30m x 0.32mm) was used. Allo-tetrahydroDOC was used as internal standard. A standard curve was constructed by SIM of m/z 457 using beta-cortol as a standard and correcting for the relative abundance of the ion at 457 in beta-cortol (0.231%). To validate the method, beta-cortolone, which produces the ion at m/z 449 was assayed using both beta-cortol and beta-cortolone as standards.

Results. Urinary beta-cortolone excretion rates in patients and normal subjects (n=19) quantitated using authentic (beta-cortolone) standard and (beta-cortol) standard were similar (R(super)2: 0.998, p<0.001). For 18-OH-THA, quantitation using the alternative standard (beta-cortol), the within and between batch coefficients of variation were 12.1 and 12.5% respectively. The lower limit of detection was 2.4 micrograms per litre. In 32 healthy adult volunteers (18 male, 14 female; age: 18-65 years), excretion rates ranged from 20 to 204 micrograms per 24hours. In 3 patients with aldosterone-secreting adenomas, the values were 1868, 963 and 253 micrograms per 24hours. In 2 patients with adrenal hyperplasia and high plasma aldosterone, the values were 2463 and 1266 micrograms per 24hours. These results are in good agreement with the few results previously published (2).

Conclusion. The use of alternative standards may provide a useful alternative to authentic standards where the latter are not readily available.

References.

1- C.H.L. Shackleton. Endocr Rev. 6 (1985) 441.

2- S. Ulick. J.Clin. Endocrinol. Metab. 43 (1976) 92.

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