Lipids

, Volume 42, Issue 8, pp 765–776

Liquid Chromatography–Mass Spectrometric Analysis of Lipids Present in Human Meibomian Gland Secretions

  • Igor A. Butovich
  • Eduardo Uchiyama
  • Mario A. Di Pascuale
  • James P. McCulley
Original Article

Abstract

The purpose of the study was to qualitatively characterize the major lipid species present in human meibomian gland secretions (MGS) by means of high-performance liquid chromatography with atmospheric pressure ionization mass spectrometric detection of the analytes (NP HPLC-MS). Two different NP HPLC-MS methods have been developed to analyze lipid species that were expected to be present in MGS. The first method was optimized for the analysis of relatively nonpolar lipids [wax esters (WE), di- and triacyl glycerols (DAG and TAG), cholesterol (Chl) and its esters (Chl-E), and ceramides (Cer)], while the second method was designed to separate and detect phospholipids. The major lipid species in MGS were found to be WE, Chl-E, and TAG. A minor amount of free Chl (less then 0.5% of the Chl-E fraction) was detected in MGS. No appreciable amounts of DAG and Cer were found in MGS. The second NP HPLC-MS method, capable of analyzing model mixtures of authentic phospholipids (e.g. phosphatidylglycerol, phosphatidylethanolamine, phosphatidic acid, phosphatidylinositol, phosphatidylserine, phosphatidylcholine, and sphingomyelin) in submicrogram/mL concentrations, showed little or no presence of these species in the MGS samples. These observations suggest that MGS are a major source of the nonpolar lipids of the WE and Chl-E families for the tear film lipid layer (TFLL), but not of the previously reported phospholipid components of the TFLL.

Keywords

Lipids HPLC Mass spectrometry Tear film lipid layer Human meibomian gland 

Abbreviations

APCI

Atmospheric pressure chemical ionization

API

Atmospheric pressure ionization

Cer

Ceramide

Chl

Cholesterol

Chl-E

Cholesteryl ester

Chl-O

Cholesteryl oleate

DA

Diarachidoylglycerol

DAG

Diacyl glycerol

DES

Dry eye syndrome

ESI

Electrospray ionization

GC

Gas chromatography

HP

n-Hexane/propan-2-ol (95:5, by vol) solvent mixture

HPA

n-Hexane/propan-2-ol/acetic acid (95:5:0.1, by vol) solvent mixture

HPLC

High-performance liquid chromatography

IS-CID

Ion source collision induced dissociation

MGS

Meibomian gland secretions

MS

Mass spectrometry

NMR

Nuclear magnetic resonance spectroscopy

NP-HPLC

Normal-phase HPLC

C16:0/C18:1-PA

1-Palmitoyl-2-oleoyl-phosphatidic acid

C16:0/C18:1-PC

1-Palmitoyl-2-oleoyl-phosphatidylcholine

C16:0/C16:0-PE

1,2-Dipalmitoyl-phosphatidylethanolamine

C16:0/C18:1-PG

1-Palmitoyl-2-oleoyl-phosphatidylglycerol

C18:0/C20:4-PI

1-Stearoyl-2-arachidonoyl-phosphatidylinositol

PL

Phospholipid(s)

C18:0/C18:1-PS

1-Stearoyl-2-oleoyl-phosphatidylserine

RT

Retention time (min)

C18:0-SM

C18:0-sphingomyelin

SS

Stearyl stearate

TAG

Triacyl glycerol

TFLL

Tear film lipid layer

TP

Tripalmitoylglycerol

WE

Wax ester

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Copyright information

© AOCS 2007

Authors and Affiliations

  • Igor A. Butovich
    • 1
  • Eduardo Uchiyama
    • 1
  • Mario A. Di Pascuale
    • 1
  • James P. McCulley
    • 1
  1. 1.Department of OphthalmologyUniversity of Texas Southwestern Medical Center at DallasDallasUSA

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