Quantitation of In Vitro α-1 Adrenergic Receptor Antagonist Binding Capacity to Biologic Melanin Using Tandem Mass Spectrometry. (December 2013)

• Record Type: Journal Article
• Authors: Gaynes, Jeffrey S.; Micic, Cedomir; Gaynes, Bruce I.; Borgia, Jeffrey A.
• Is Part Of: Current eye research. Volume 38:Number 12(2013:Dec.)
• Page Start: 1214
• Page End: 1220
• Publication Date: 2013-12
• Subjects: Ophthalmology -- Periodicals; Eye -- Diseases -- Periodicals; Ophthalmology -- Periodicals; 573.88
• DOI: 10.3109/02713683.2013.822894
• Languages: English
• ISSNs: 0271-3683

Abstract

<abstract> <title>Abstract</title> <p> <italic>Purpose</italic>: The purpose of this study was to develop methods to allow evaluation of the binding characteristics for a series of α-1 antagonists to biologically-derived melanin.</p> <p> <italic>Methods</italic>: Fresh bovine globes were used to obtain iridal and choroid/retinal pigment epithelial (CRPE) derived melanin. Binding characteristics of chloroquine, tamsulosin and doxazosin were then evaluated <italic>in vitro</italic> using tandem mass spectroscopy.</p> <p> <italic>Results</italic>: Tandem mass spectrometry-based assays were developed for three α-1 antagonists that provided linear assay ranges which spanned (minimally) 0.01–10 µg/mL, while exhibiting excellent inter-assay precision and accuracy. When applied to the evaluation of binding characteristics for iridal melanin, mean chloroquine and tamsulosin fractions were found to be 41.9 ± 14.2 pmoles mg<sup>−1</sup> and 25.34 ± 6.186 pmoles mg<sup>−1</sup>, respectively. Mean iridal doxazosin binding was found to be 6.36 ± 2.19 pmoles mg<sup>−1</sup>. Interestingly, mean levels of tamsulosin, but not doxazosin found bound to choroid/CRPE derived melanin approached that of chloroquine (27.91 µg/mL, 25.68 µg/mL and 5.94 µg/mL for chloroquine, tamsulosin and doxazosin, respectively). One way ANOVA for binding affinity for chloroquine, tamsulosin and doxazosin was statistically significant for both iridal and CRPE-derived melanin (<italic>p</italic> = 0.0012 and 0.0023), respectively. A Bonferroni <italic>post-hoc</italic> analysis demonstrated a statistically significant difference in the amount of binding between tamsulosin, doxazosin and chloroquine to iridal but not CRPE derived melanin (<italic>p</italic> < 0.05).</p> <p> <italic>Conclusions</italic>: Tamsulosin appears to demonstrate melanin binding affinity which approaches chloroquine and exceeds doxazosin for both iridal and CRPE-derived bovine melanin.</p> </abstract>