Producing Deuterated Oleic Acid at the National Deuteration Facility

Authors

Tamim Darwish, Emily Luks, and Marie Gillon

Introduction

Oleic acid is a mono-unsaturated fatty acid, with a cis-orientation about its double bond (Figure 1), and is a major component in common vegetable oils such as olive, peanut, grape seed and sesame oils.

It is also the most abundant fatty acid present in many animal fats (chicken, pig, turkey and human). Oleic acid is also emitted by the decaying corpses of certain insects such as bees or ants, so that live workers know to remove them from the hive.

At the molecular level, oleic acid also forms an unsaturated tail component in many of the phospholipid molecules (such as DOPC - Figure 2) that are fundamental to the structure and function of cellular membranes. Biophysical investigations into how cellular membranes function are of vital importance in understanding the mechanisms of neurodegenerative diseases (such as Parkinson’s and Alzheimer’s), as well as the action of bacterial toxins (e.g. diphtheria, cholera, pneumonia, and staphylococcus) on our cells.
 

Deuteration of phospholipids or other long-chain fatty acids is an essential first step in many neutron-scattering experiments; particularly those associated with Small-Angle Scattering (Quokka) or Reflectometry (Platypus). When studying surfactants or lipids at the air-water interface, a non-deuterated hydrocarbon chain has a Scattering Length Density (SLD) almost identical to that of air and it is essentially invisible to neutrons.

However, when deuterated, these chains have a large SLD and stand-out very clearly from both air and the water subphase. In other studies, one may wish to observe the interaction between other hydrocarbon-based chemicals or biological molecules with lipids or surfactants. Again, there is a lack of contrast which can be overcome by deuterating of the tail component of these lipids or fatty acids.

Deuteration is typically achieved by reacting the hydrocarbon chains of the corresponding saturated fatty acids under hydrothermal conditions with D2O and a metal catalyst in a Parr reactor (~220°C, 22 atmospheres). This is the usual method of producing deuterated precursors for the synthesis of phospholipids such as DPPC, where the tails are fully saturated.

However, the Parr reactor method is not suitable for direct deuteration of oleic acid, as it leads to hydrogenation of the C=C bond (forming stearic acid), or isomerisation leading to formation of the trans isomer of oleic acid (elaidic acid). As a consequence, deuterated oleic acid is not commercially available; nevertheless, in the light of its importance in scientific studies using neutrons we have devised a method to synthesize it by constructing the molecule from deuterated precursors, using multi-step organic reactions.

A synthetic route to deuterated oleic acid makes use of the deuterated fatty acid precursors that we can produce in our Parr reactors on a large scale. In order to preserve the high level of deuteration, as well as the cis-orientation of the double bond, nine separate chemical reactions (with associated purification and characterisation steps) were required to produce deuterated oleic acid.

The full reaction sequence was a very labor intensive process, requiring the attention of several members of the National Deuteration Facility (Figure 4), but the benefit is that we have now successfully produced gram-scale quantities of deuterated oleic acid.

The immediate fate of this first batch of deuterated oleic acid is to be used in the study of lyotropic liquid crystal nanoparticles (cubosomes) using Small Angle Neutron Scattering on Quokka in a collaboration between researchers at ANSTO and Monash University.