Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS) Assay for Simultaneous Measurement of Salivary Testosterone and Cortisol in Healthy Men for Utilization in the Diagnosis of Late-onset Hypogonadism in Males

Abstract

Abstract. it is well known that late-onset hypogonadism in males can cause a variety of symptoms, and the differential diagnosis is relatively difficult, including psychological disorders, stress, and mood disturbances. The level of serum cortisol can be measured to reflect a patient's level of stress. Salivary hormones facilitate the evaluation of physiological hormonal actions based on free hormone assay. For the simultaneous measurement of testosterone and cortisol levels in saliva, we validate a sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay. Concerning accuracy and precision, the lower limit of quantification of salivary testosterone and cortisol were established as 5 and 10 pg/mL, respectively. Testosterone and cortisol in saliva is stable for 2 days, 14 days, and 28 days at room temperature, refrigeration and frozen, respectively. Freezing and thawing for 3 cycles and stimulation of salivation with gum chewing do not alter the measured values of testosterone and cortisol. Total, bioavailable, and free serum testosterone showed slight diurnal changes, but total and bioavailable serum cortisol showed marked diurnal changes. Salivary testosterone levels negatively correlate with age, regardless of the time of saliva collection (r=0.64, p<0.05). However, there is no relationship between salivary cortisol and age (r=0033, p>0.05). LC-MS/MS allows rapid, simultaneous, sensitive, and accurate quantification of testosterone and cortisol in saliva for the diagnosis late-onset hypogonadism or other hormone related disease. 2]. The differences may be associated with molecular structure variations in water solubility, differences in binding affinity to sex hormone-binding globulin (shBg) in serum, or the serum free hormone ratio [3]. However, the mechanism by which hormones are secreted in saliva via blood remains to be clarified. Autoradiograms showed free T was secreted in saliva without binding to SHBG [4]. Recent studies reported t and F in saliva were useful for evaluating bioavailability and accurately reflected serum hormone activity [1,3,[5][6][7][8]. Some studies indicated the measurement of salivary hormone levels facilitated the evaluation of precise physiologic actions based on free hormone activity, which could not be confirmed based on total blood hormone levels [1,3,6,9]. therefore, salivary hormone measurements have re-

Assessment of salivary free cortisol levels by liquid chromatography with tandem mass spectrometry (LC-MS/MS) in patients treated with mitotane

“…[12][13][14][15] The purpose of our study was to assess SC during the follow-up of patients treated with mitotane using two different methods: the electrochemiluminescence immunoassay (ECLIA), a widely employed method in the clinical laboratory, and high-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (LC-MS/MS), considered as the reference method.…”

2128 High Testosterone Levels in Prostate Tissue Obtained by Needle Biopsy Correlate With Poor Prognosis Factors in Prostate Cancer Patients

“…The method was validated to ensure that the result was within the 20% range for accuracy and precision [7]. The determination limit of the method was 0.5 pg/shot for T and 1 pg/shot for DHT.…”

“…LC-MS/MS can be used to measure androgen concentrations in tissue samples as small as those obtained by a single needle biopsy (approximately 3 mg), and the latest LC-MS/MS technique is more than 10 times as sensitive as the RIAs used in the past, especially in the lower concentration range [7].…”

Salivary Hormone Measurement Using LC/MS/MS: Specific and Patient-Friendly Tool for Assessment of Endocrine Function

“…[34][35][36] LOH is defined as a clinical and biochemical syndrome associated with advancing age and is characterized by typical symptoms (i.e., decrease in intellectual activity and cognitive functions, depressed mood and irritability, sleep disturbances, decrease in muscle volume and strength, increase in visceral fat, decrease in body hair and skin alterations, decreased bone mineral density as well as decreased libido) and a deficiency in serum testosterone levels. 37) Currently, serum free testosterone is being used for the diagnosis of LOH.…”

“…An LC/positive ESI-MS/MS assay developed by Matsui et al enabled the simultaneous determination of testosterone and cortisol without derivatization. 36) As described above, LOH causes a variety of symptoms and the differential diagnosis is relatively difficult, including psychological disorders, stress, and mood disturbances. Because the salivary cortisol level is a good marker of stress, the simultaneous determination of testosterone and cortisol is expected to be useful for the differential diagnosis of LOH and stress-induced disorders.…”