Abstract
Aqueous humor (AH) is a transparent fluid which fills the anterior and posterior chambers of the eye. It supplies nutrients and removes metabolic waste from avascular tissues in the eye. Proper homeostasis of AH is required to maintain adequate intraocular pressure as well as optical and refractive properties of the eye. Application of metabolomics to study human AH may improve knowledge about the molecular mechanisms of eye diseases. Until now, global analysis of metabolites in AH has been mainly performed using NMR. Among the analytical platforms used in metabolomics, LC-MS allows for the highest metabolome coverage. The aim of this study was to develop a method for extraction and analysis of AH metabolites by LC-QTOF-MS. Different protocols for AH preparation were tested. The best results were obtained when one volume of AH was mixed with one volume of methanol : ethanol (1 : 1). In the final method, 2 µL of extracted sample was analyzed by LC-QTOF-MS. The method allowed for reproducible measurement of over 1000 metabolic features. Almost 250 metabolites were identified in AH and assigned to 47 metabolic pathways. This method is suitable to study the potential role of amino acids, lipids, oxidative stress, or microbial metabolites in development of ocular diseases.
Highlights
Metabolomics aims to identify andquantify the small molecule metabolites present in a studied sample [1]
The purpose of this study was to develop a method for metabolic fingerprinting of human Aqueous humor (AH)
AH is to some extent similar to blood plasma as it contains plasma proteins and metabolites which are filtered to AH through fenestrated capillaries of the ciliary body stroma via the iris root [14]
Summary
Metabolomics aims to identify and (semi-)quantify the small molecule metabolites present in a studied sample [1]. This approach has been extensively applied in biomedical research to find disease biomarkers [1], study therapeutic effects of drugs or natural substances with potential therapeutic capabilities [2], or explore metabolic pathways perturbed by a particular disease or condition [3]. Regarding the samples of mammalian origin, blood (serum/plasma) and urine are most commonly sampled for metabolomics. These types of biological fluids are especially important in the field of biomarkers discovery [1]. Among other types of mammalian samples metabolomics studies on cerebrospinal [8] or amniotic [9] fluids, exhaled breath condensate [10], stool, or saliva [11] samples were performed
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