Parallel diurnal fluctuation of testosterone, androstenedione, dehydroepiandrosterone and 17OHprogesterone as assessed in serum and saliva: validation of a novel liquid chromatography-tandem mass spectrometry method for salivary steroid profiling.
BACKGROUND
Salivary androgen testing represents a valuable source of biological information. However, the proper measurement of such low levels is challenging for direct immunoassays, lacking adequate accuracy. In the last few years, many conflicting findings reporting low correlation with the serum counterparts have hampered the clinical application of salivary androgen testing. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) makes it possible to overcome previous analytical limits, providing new insights in endocrinology practice.


METHODS
Salivary testosterone (T), androstenedione (A), dehydroepiandrosterone (DHEA) and 17OHprogesterone (17OHP) were extracted from 500µL of saliva, separated in 9.5 min LC-gradient and detected by positive electrospray ionization - multiple reaction monitoring. The diurnal variation of salivary and serum androgens was described by a four paired collection protocol (8 am, 12 am, 4 pm and 8 pm) in 19 healthy subjects.


RESULTS
The assay allowed the quantitation of T, A, DHEA and 17OHP down to 3.40, 6.81, 271.0 and 23.7 pmol/L, respectively, with accuracy between 83.0 and 106.1% for all analytes. A parallel diurnal rhythm in saliva and serum was observed for all androgens, with values decreasing from the morning to the evening time points. Salivary androgen levels revealed a high linear correlation with serum counterparts in both sexes (T: R>0.85; A: R>0.90; DHEA: R>0.73 and 17OHP: R>0.89; p<0.0001 for all).


CONCLUSIONS
Our LC-MS/MS method allowed a sensitive evaluation of androgen salivary levels and represents an optimal technique to explore the relevance of a comprehensive androgen profile as measured in saliva for the study of androgen secretion modulation and activity in physiologic and pathologic states.
['Mezzullo|Marco|M|http://orcid.org/0000-0001-6748-3041', 'Fazzini|Alessia|A|', 'Gambineri|Alessandra|A|', 'Di Dalmazi|Guido|G|', 'Mazza|Roberta|R|', 'Pelusi|Carla|C|', 'Vicennati|Valentina|V|', 'Pasquali|Renato|R|', 'Pagotto|Uberto|U|', 'Fanelli|Flaminia|F|http://orcid.org/0000-0003-2601-2694']
D019326:17-alpha-Hydroxyprogesterone D000328:Adult D000368:Aged D000735:Androstenedione D001774:Blood Chemical Analysis D002853:Chromatography, Liquid D002940:Circadian Rhythm D003687:Dehydroepiandrosterone D005260:Female D006801:Humans D008297:Male D008875:Middle Aged D012463:Saliva D053719:Tandem Mass Spectrometry D013739:Testosterone
testosterone androstenedione 17OHProgesterone saliva liquid chromatography-tandem mass spectrometry dehydroepiandrosterone (DHEA)