Abstract
Purpose :
Analysis of sex hormones in tear fluid will aid understanding of their role in regulation of tear production and in dry eye disease. Levels of sex hormones in tears are unknown due to low tear volumes available and a lack of sufficiently sensitive quantification methods. This pilot study applied an ultrasensitive LC-MS method to estimate androgens and oestrogens in tears of pre- and post-menopausal women.
Methods :
An ultrasensitive LC-MS method previously developed for serum sex hormones was applied. The accuracy of this method for tears was assessed by absolute recovery from pooled basal tears of pre-menopausal women spiked with three levels of known amounts of sex steroids. Flush tears (where 60 µL saline was applied to the ocular surface, collected using a microcapillary tube and samples pooled from both eyes/participant) from pre- and post-menopausal women (n=2/category) were subsequently analysed for testosterone (T), estradiol (E2) and progesterone (P4) concentration on an API-5000 triple-quadruple mass spectrometer equipped with an atmospheric pressure photoionization source. Recovery tests were run in six replicates and other analyses were run in duplicate.
Results :
Excellent absolute recoveries were observed for all steroids analysed at the concentrations tested, with accuracy ranging from 93.7% (P4) to 99.3% (T) for basal tears. Concentrations of E2 were 2.90±1.01 pg/mL in flush tears of post-menopausal women and 8.40±1.70 pg/mL in pre-menopausal women. P4 was 0.30±0.07 ng/mL in flush tears of pre-menopausal women but was not quantifiable in all tear samples from post-menopausal women. T was not detectable in any of the flush tear samples.
Conclusions :
The ultrasensitive LC-MS method showed high accuracy for analysis of sex hormones in basal tears. The method effectively quantified E2 and P4 in flush tears of pre- and post-menopausal women. Further work will explore this method for analysis of sex hormones in flush and saline-diluted basal tears to tackle the limitations of tear sample volume
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.