In this interview, News Medical speaks with Professor Brian Keevil about the growing role of salivary cortisol and salivary cortisone testing in the diagnosis and monitoring of adrenal disorders, including Cushing's syndrome and adrenal insufficiency. He discusses how non-invasive saliva-based testing, combined with LC-MS/MS analysis, could improve diagnostic accuracy, reduce the need for hospital visits, and support earlier intervention for patients with endocrine disorders.
Can you please introduce yourself and your role at Manchester University NHS Foundation Trust?
My name is Professor Brian Keevil, and I am the Clinical Lead for Biochemistry at Manchester University NHS Foundation Trust. My work focuses on the development and implementation of advanced mass spectrometry methods for clinical diagnostics, particularly in endocrinology. Over the last 15 years, much of our research has centered on improving the measurement of steroid hormones, including cortisol and cortisone, to support the diagnosis and monitoring of adrenal disorders such as Cushing's syndrome and adrenal insufficiency.
What sparked your interest in using salivary cortisol and cortisone for investigating adrenal disease?
We were interested in finding a simpler and more patient-friendly alternative to traditional blood-based testing. Saliva is very easy to collect, convenient for patients, and can often be collected at home. Unlike blood sampling, it avoids venepuncture and reduces the burden on both patients and healthcare services.
From an analytical perspective, saliva contains the free fraction of cortisol, which is the biologically active component. This makes it particularly useful when assessing adrenal function. Over time, we found that measuring salivary cortisone offered several advantages over salivary cortisol, particularly in terms of sensitivity and correlation with serum cortisol concentrations.
What are the main considerations when collecting saliva samples for cortisol and cortisone testing?
Saliva collection is straightforward, but it is important to follow correct procedures. Samples can be collected using either passive drool methods or specialized swabs such as Salivette or SalivaBio devices. Fortunately, cortisol and cortisone are largely unaffected by the collection device used.
Patients should avoid eating, drinking, or brushing their teeth for at least 30 minutes before collection because blood contamination from gum irritation can affect results. Dehydration can also make collection difficult, and certain conditions, such as Sjögren's syndrome, may prevent adequate saliva production altogether.
The good news is that cortisol and cortisone are very stable in saliva. Samples can remain stable at room temperature for several days, although refrigeration or freezing is preferable if there will be any delay before analysis.
Why has salivary cortisone emerged as a particularly valuable biomarker compared with salivary cortisol?
What we are really interested in clinically is free cortisol, which represents only around 5% of circulating cortisol because the majority is bound to proteins. When free cortisol enters the salivary gland, much of it is converted into cortisone by the enzyme 11β-hydroxysteroid dehydrogenase type 2.
As a result, salivary cortisone concentrations are around four times higher than salivary cortisol concentrations, making them easier to measure accurately. More importantly, salivary cortisone shows a much more linear relationship with serum cortisol levels. That means it more closely reflects physiological cortisol exposure and provides a more reliable marker across a wider concentration range.
How could salivary cortisone improve the diagnosis of adrenal insufficiency?
Adrenal insufficiency is a potentially life-threatening condition that is often underdiagnosed because symptoms are nonspecific and existing screening tests can be inconvenient or resource-intensive.
Traditionally, diagnosis relies on the ACTH stimulation test, also known as the Short Synacthen Test. While effective, this requires hospital attendance and specialist staff. Our work has shown that waking salivary cortisone collected at home can act as an effective first-line screening test.
We have demonstrated that the test is safe, practical, and capable of reducing the number of patients requiring more invasive investigations. Several UK hospitals are now implementing or evaluating this approach, and Manchester University NHS Foundation Trust has already begun routine use of the test.
How effective is salivary cortisone in diagnosing Cushing's syndrome?
Cushing's syndrome remains one of the most challenging endocrine disorders to diagnose because there is no single definitive test. Current investigations include urinary free cortisol measurements, late-night salivary cortisol, and overnight dexamethasone suppression testing.
Our studies have shown that late-night salivary cortisone performs extremely well, particularly in patients with pituitary-dependent Cushing's disease. In one tertiary referral study, late-night salivary cortisone was the single best-performing test, achieving sensitivity of 95% and specificity of 100% using an optimized cutoff value. It outperformed both urinary free cortisol and standard late-night salivary cortisol measurements.
What have you learned about using salivary cortisone alongside the overnight dexamethasone suppression test?
The overnight dexamethasone suppression test is widely used because dexamethasone should suppress cortisol production in healthy individuals. We investigated whether saliva samples could replace serum measurements in this setting.
The results were very encouraging. We found strong correlations between serum cortisol and post-dexamethasone salivary cortisone concentrations. Using a salivary cortisone cutoff of less than 2.7 nmol/L produced excellent diagnostic performance, with sensitivity around 85% and specificity around 92% for detecting cortisol excess.
Importantly, saliva-based testing removes the need for patients to attend the hospital the morning after taking dexamethasone, making the process significantly more convenient.
Your research has also explored salivary dexamethasone metabolites. Why is that important?
One challenge with dexamethasone suppression testing is confirming that patients have actually absorbed the medication correctly. In serum testing, we often measure dexamethasone directly for this reason.
Direct measurement of salivary dexamethasone has not performed particularly well. However, we found that measuring salivary 11-dehydrodexamethasone, a metabolite produced through similar enzymatic pathways to cortisone, provides much better correlation with serum dexamethasone concentrations.
This could become a valuable addition to future saliva-based diagnostic pathways and may help identify patients whose suppression tests fail because of altered dexamethasone metabolism or absorption.
How might salivary cortisone testing help clinicians identify milder forms of cortisol excess?
One of the areas generating considerable interest is mild autonomous cortisol secretion, which can be difficult to identify using conventional approaches.
Recent work suggests that late-night salivary cortisone may offer excellent discrimination between affected patients and controls. In a recent study, we observed strong sensitivity and specificity using carefully selected thresholds, suggesting that salivary cortisone could become an important screening tool for identifying subtle cortisol excess before more severe disease develops.
What future developments do you see for salivary cortisone testing in endocrine medicine?
We are currently involved in multicenter studies across the UK and Germany investigating cyclical Cushing's syndrome, a condition where cortisol excess fluctuates over time. By collecting repeated late-night saliva samples over several weeks, we hope to better understand disease patterns that are often missed using conventional testing.
More broadly, I believe salivary cortisone will continue to gain traction as a routine outpatient and home-based test. It offers excellent diagnostic performance, is highly convenient for patients, and has the potential to reduce healthcare costs while improving access to endocrine testing. As more centers adopt LC-MS/MS methodologies, I expect saliva-based steroid testing to become an increasingly important part of clinical practice.
About Professor Brian Keevil
Professor Brian Keevil is a Consultant Clinical Scientist at Manchester University NHS Foundation Trust. He is Clinical Lead for the Clinical Biochemistry Department at Wythenshawe hospital and director of the clinical mass spectrometry unit.
His research interests involve the development and introduction of novel mass spectrometry methods for the measurement of therapeutic drugs and steroids. His laboratory has pioneered the use of these techniques many of which are used to underpin routine clinical services and collaborative international research projects.
Steroids have been a particular interest, and the laboratory performs a wide range of steroid assays in blood urine and saliva samples. These methods are being used to refine existing endocrine protocols and to explore the use of newer novel steroids such as the 11 oxygenated steroids in the diagnosis and management of endocrine conditions. He is the recipient of a UKRI scholars grant to investigate the use of salivary cortisone in the investigation of adrenal disease.
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