Nuclear Magnetic Resonance (NMR) spectroscopy is one of the most powerful tools for elucidating the structural and dynamic features of biomolecules and their biochemical interactions. However, the inherently low sensitivity of NMR remains a significant limitation, primarily due to the low population differences (spin polarization) between spin states at thermal equilibrium. To overcome this challenge, a number of complementary hyperpolarization techniques were developed, which, by physical or chemical techniques, create spin-polarization far away from thermal equilibrium and boost the NMR signals by several orders of magnitude. Parahydrogen Induced Polarization (PHIP) and its reversible variant SABRE (Signal Amplification By Reversible Exchange) are among the most versatile and cost-efficient hyperpolarization techniques for NMR signal enhancement in solution NMR.

The talk will first give a short introduction to PHIP and SABRE, followed by an overview of our special PHIP set-ups. Then some examples from our recent work on bioactive or biological molecules will be presented. The first example shows the results of our studies on a bioactive derivative of sunflower trypsin inhibitor-1 (SFTI-1), which inhibits matriptase, a colon cancer-related enzyme. The PHIP activity of the inhibitor was achieved by labeling the tetradecapeptide with O-propargyl-L-tyrosine. By using a carefully optimized automated PHIP setup in 1D PHIP experiments, an amplification of up to approximately 1200 was found compared to normal NMR. This enormous amplification factor enabled ultra-fast single-scan detection of 2D TOCSY spectra of micromole solutions of the PHIP-labeled inhibitor. We then present recent results on the hyperpolarization of fumarate and eptifibatide, a platelet aggregation inhibitor derived from the venom of certain rattlesnakes, and octreotide, a somatostatin analogue used in the diagnosis and treatment of various cancers. While the previous examples used rather small target molecules, the two remaining examples show the application of PHIP to the trypsin inhibitor (MCoTI) from Momordica Cochinchinensis a disulfide-rich mini-protein and the 236 kD biopolymer poly-γ-(4-propargyloxy)-benzyl-L-glutamate.