Phospholipids are the primary structural constituents of biological membranes, which
have been reported to exert various functions in biological systems. However, the
phospholipids can't be well prepared by means of traditional Bligh & Dyer and Folch
methods, because a lot of matrix would be co-extracted leading to the signal
suppression of phospholipids in the mass spectrum. Therefore, an efficient analytical
strategy is highly demanding. The aim of this project was to 1) establish and optimize
solid phase based strategies for extracting phospholipids from foods; 2) establish MS
based detecting methods for qualitative and quantitative analysis of phospholipids.
Hydrophilic interaction chromatography (HILIC) is a powerful tool in separating
phospholipid classes. In the first part of this study, HILIC (silica derivatized with
1,2-dihydroxypropane, SiO2/diol) was used as a new solid-phase extraction (SPE)
sorbent for the selective extraction of phospholipids. Matrix-assisted laser
desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) was
explored for fast profiling of phospholipids in salmon, which is a delicious food but
easily susceptible to spoilage by microorganism. By a protocol developed, a total
number of 30 phospholipids and 16 lysophospholipids were qualitatively and
quantitatively detected. The effect of Pseudomonas fluorescens on the changes of
phospholipids in salmon fillet during shelf life period was measured. It showed that
the phospholipids in the m/z range from 700 to 900 were degraded to
lysophospholipids by phospholipase, along with the extension of storage, indicating
that salmon fillet was easily rotten once contaminated with P. fluorescens.
Although HILIC performed well in extracting phospholipids, the SiO2/diol
sorbent is 10 times more expensive in comparison with titanium dioxide (TiO2).
Hence, in the second part of this study, TiO2, which has been reported to be efficient
in selectively extracting phosphorous compounds, was chosen for purifying
phospholipids. Olive oil and almond were analyzed as representative food materials,
because they are rich in phospholipids. A matrix solid-phase dispersion (MSPD)
procedure using TiO2 nanoparticle (NP) as sorbent combined with a MALDI-TOF/MS
method was developed for selective extraction, fast visualization and analysis of
phospholipids. The results proved that clean MS spectrum could be obtained when
TiO2 NP was utilized as MSPD sorbent, with acidified water (pH 5) for washing the
matrix and enhancing the binding ability, followed by chloroform and methanol (1/2,
v/v) as eluting solvent. Based on this approach, phospholipids could be selectively
separated from olive and almonds samples. The ratio of m/z 833.6/835.6 and the ion
of m/z 821.6 could be used as potential markers for differentiating the geographical
origins of almonds.
The MSPD cartridge is disposable and can't be recycled. In the third part of this
study, TiO2/SiO2 core-shell composite was synthesized, and used as SPE sorbent for
extracting phospholipids from shrimp waste (Litopenaeus vannamei). The
performance of SPE protocol was evaluated, and the optimum conditions were found
to be: pH 5 of loading solvent, 10% aqueous methanol as washing solvent and 1.0 mL
of chloroform/methanol (1/2, v/v) as eluting solvents. The results indicated that,
compared with previously published methods, this strategy was cost-effective and
efficient. Meanwhile, it was found that phospholipids were abundant in shrimp waste,
and the major phospholipid species of each class were PC 16:0/22:6 (480.7 μg∙g-1),
aPE 18:1/20:5 (378.0 μg∙g-1), PI 18:0/20:5 (45.2 μg∙g-1) and PS 16:1/22:5 (47.6
μg∙g-1). The successful application of this strategy paves the way for full utilization of
traditionally discarded shrimp wastes.
Graphene is a versatile material, and integrating graphene with other inorganic
materials to fabricate composites or hybrids is the focus of the applications. For this
reason, graphene and titanium dioxide (G/TiO2) composite was synthesized and used
as SPE sorbent for the selective extraction of phospholipids from avocado (Persea
americana Mill). It showed that non-phospholipid such as diglycerides, triglycerides
and their fragments were removed, and ions intensities of m/z 520.3, 542.3, 544.3
were increased more than two times. As a conclusion, the G/TiO2 was proved to be a
promising sorbent lipidomics study of avocado samples.
In summary, this study provides several protocols (combinations of solid phase
based sample preparation and MS based analytical methods) for selective extracting
and detecting phospholipids from complex food samples. The sorbents used in this
study, including SiO2/diol, TiO2, TiO2/SiO2 and G/TiO2, performed well and are
shown to be excellent in lipidomics study of different foods.
| Date of Award | 2 Oct 2015 |
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| Original language | English |
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| Awarding Institution | - City University of Hong Kong
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| Supervisor | Hon Yeung CHEUNG (Supervisor) |
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