Abstract:
Immune responses to infection by the human immunodeficiency virus (HIV) and the use of highly active
antiretroviral therapy (HAART) to treat HIV infection, contributes to metabolic irregularities in the host.
Current methods for the extraction and identification of metabolites in biofluids generally make use of
laborious, time-consuming protocols. Here, 96-well Ostro™ plates and filtration under positive pressure was
used to facilitate the simultaneous, reproducible extraction of metabolites from multiple serum samples which
were then analyzed by ultra-performance liquid chromatography mass spectrometry (UPLC-MS). The easy to
use solid phase extraction (SPE) protocol eliminated numerous potential contaminants while the UPLC-MS
detection of metabolites produced visibly different chromatograms for HIV negative (n=16), HIV+ (n=13) and
HIV+HAART+ (n=15) serum samples. Linear discriminant analysis (LDA) amplified these differences,
classified the groups with 100% accuracy and identified biomarkers explaining the greatest variances between
the groups. The 21 metabolites altered by HIV and/or HAART primarily represented those linked to lipid and
energy pathways which is where known metabolic changes associated with HIV infection occur. This work
demonstrated for the first time that OstroTM plates and UPLC-MS metabonomics was able to successfully
identify distinct differences between the experimental groups and detected metabolites related to HAART and
other drugs used in the treatment of HIV-associated conditions. The findings of this approach suggests a
possible role for this methodology in disease prognosis as well as in the monitoring of treatment success or
failure and linking treatment to metabolic complications.
