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Towards a better understanding of uraemic molecules

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Nouri Koupaei, M (2013) Towards a better understanding of uraemic molecules. Research Master thesis, University of Tasmania.

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Abstract

Currently, there is no‐cure or pre‐diagnosis test for patients suffering
from Chronic Kidney Disease (CKD). Uremic toxins which normally excreted
into the urine by healthy kidneys accumulate in the body due to kidney
malfunction causing complications such as higher risk of cardiovascular
disease. These complications associated with CKD leading to the high
mortality rate among the patients. Although the removal of the uraemic
solutes such as urea and creatinine can be achieved by dialysis but it does
not affect the survival rate. Therefore, it is suggested that there are some
other toxins that cannot be removed by hemodialysis causing the symptoms
of CKD. Analysis of serum samples of patients suffering from CKD before
and after dialysis compared to samples collected from healthy volunteers can
thus give more insight into the effect of uraemic solutes as those remaining
after dialysis can lead to the death. In addition, monitoring the changes in
metabolites after dialysis can be very helpful to evaluate dialysis membrane
efficiency in removal of uraemic toxins.
Analyses of uraemic molecules in serum sample of a group of patients
suffering from CKD were performed with three analytical methods
including capillary electrophoresis‐mass spectrometry (CE‐MS), gas
chromatography‐mass spectrometry (GC‐MS) and liquid chromatographymass
spectrometry (LC‐MS), and the results compared with a healthy
(control) group. Both targeted and non‐targeted (global profiling) studies
were aimed. Unlike GC‐MS and LC‐MS which proved to be suitable for the
analysis of respectively polar and non‐polar metabolites, CE‐MS practically failed in providing reproducible results to be compared to the other two
techniques.
Targeted study of some of the known uraemic toxins using GC‐MS
was performed employing the available individual standards following
suitable derivatisation procedures. On the other hand, higher numbers of
known uraemic molecules were identified in LC‐MS upon careful
monitoring of MS features such as exact masses of molecular ions and
adducts in both positive and negative ion‐modes using available data bases
such as METLIN library and Human Metabolome Database. Comparison of
the results shows that most of the compounds had significant reduction post
dialysis, which also reflects the quality of the dialysis treatment. Also unlike
water soluble metabolites, the analysis of protein bound solutes was found
less conclusive due to the complications associated with them, for example,
in finding a suitable sample preparation to be able to efficiently cleave their
bounds with proteins.
Global metabolic profiling of the results was also performed
employing XCMS platform by visualising the processed data (p‐value < 0.05)
as distribution profiles. These profiles generally show significant reduction
of detected metabolites after dialysis with metabolites had been more
efficiently removed from m/z range of 100 to 500. Accordingly, 50% of
metabolites were distributed in m/z range of 250‐550, 25% in the relatively
narrower range of 100‐250 and the rest were between m/z of 550 to 950. Also,
the majority (90%) of detected metabolites showed relative fold changes less
than 70 pre‐dialysis which reduced to about 25 after treatment. Moreover,
fold change distributions for 50% of metabolites before dialysis was approximately between 3 and 15 which reduced to less than 5 post dialysis.
On the other hand, treatment seems to have insignificant effect on fold
changes of about 10% of metabolites. As one application, such information
might be beneficial in better understanding of the performance of different
dialysis treatments.

Item Type: Thesis (Research Master)
Keywords: Chronic Kidney Disease (CKD), Uraemic Molecules, Metabolomics, Capillary Electrophoresis-Mass Spectromy (CE-MS), Gas Chromatography-Mass Spectrometry (GC-MS), Liquid Chromatography-Mass Spectrometry (LC-MS).
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Date Deposited: 02 Oct 2013 00:09
Last Modified: 15 Sep 2017 01:06
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