NMR Studies of Sperm and Seminal Plasma

The aims of this thesis were to use Nuclear Magnetic Resonance (NMR) to scan live spermatozoa in order to identify and quantify spermatozoa metabolites. The results indicate that NMR can both identify live sperm metabolites as well as quantify and monitor them in real time.

NMR has only once previously been used to study live sperm metabolism in real time. Therefore, the first half of this thesis focussed on optimising proton NMR (1H) parameters and ensuring sperm remained viable within the scanner. Six sperm metabolite peaks were identified from NMR spectra indicating the presence of lipid, lactate, arginine+spermine+leucine, citrate, glutamine and glucose/fructose. External stimuli were then used to alter sperm metabolism and the difference in metabolite spectral peaks quantified. Changes in lactate were measurable which helped support previous literature suggesting that glycolysis was a significant metabolic pathway in mammalian sperm and that NMR was capable of detecting metabolite changes in real time.

Intra ejaculate variation of metabolites was investigated in high motility, normozoospermic sperm and found to not vary signignificantly. The comparison of metabolites observed in seminal plasma, as well as low-quality and high-quality sperm was then performed between normozoospermic (n=15), oligozoospermic (n=3); and asthenozoospermic (n=5) samples. This suggested, there were no significant differences between metabolites from high and low motility spermatozoa, but glutamine was significantly different in seminal plasma between normozoospermic & oligozoospermic ejaculates and oligozoospermic & asthenozoospermic ejaculates.

Finally carbon 13 NMR was used to track labelled pyruvate in real time as it was metabolised by spermatozoa into metabolic products. Pyruvate, lactate, and bicarbonate were identified suggesting that both glycolytic and OXPHOS metabolism pathways were being utilised. CO2 was identified in longer scans further supporting sperm utilising OXPHOS alongside glycolysis. No significant difference was identified in human spermatozoa from different classes of fertility in terms of rate of 13C pyruvate consumption.