Drochaid and Oxford Biotrans collaborate on successful project
Over the past few months, Drochaid Research Services Ltd and Oxford Biotrans Ltd have been working together to use Drochaid's specialist equipment and expertise to refine the concentrations of the enzymes Oxford Biotrans use to produce their natural nootkatone product.
Recently completed, this project has produced wins for both companies, with Oxford Biotrans seeing improved efficiency and considerable knowledge gains, and Drochaid proving it can support the biotechnology sector.
Drochaid Research Services Ltd and Oxford Biotrans Ltd have worked together before in 2019, but it was in October 2020 that Oxford Biotrans approached Drochaid regarding a potential programme of work to probe the impact of substrate and enzyme concentrations on an enzymatic oxidation process. Drochaid have relevant experience in this field having previously carried out this reaction in an earlier project.
The focus of this study was the biotransformation of valencene to nootkatone using a modified patented Cytochrome P450 enzyme engineered by Oxford Biotrans.
Nootkatone is a flavour and fragrance ingredient found in low concentrations in grapefruit and oranges. Due to its low concentration, it takes 400,000 kgs of grapefruit to make 1kg of natural nootkatone. This makes EU Natural nootkatone one of the most expensive flavours in the world, costing between £3000-£4000 per kg. Valencene, however, is available from oranges at higher concentrations, reducing its market price and making it an attractive substrate.
Figure 1: Biotransformation of valencene to nootkatone
Traditional routes from valencene to nootkatone use heavy metal catalysts and peroxides and as such the product is not classed as EU Natural. The biotransformation route developed by Biotrans uses a cytochrome P450 enzyme to convert natural valencene to nootkatone (by adding an oxygen atom and removing 2 x hydrogens from valencene): under EU regulations this is regarded as an EU Natural product.
Figure2. Example of two of the 1.2L Premex reactors
Drochaid Research Services was well equipped to conduct this work using their 1.2L Premex autoclave.
The reactor is equipped with gas entraining stirring which allows for optimal mixing. The reactor is equipped with a pH probe and the capability to add base or acid using a HPLC pump to keep the pH within the desired range. The reactor is also equipped with a thermal fluid jacket to maintain a constant temperature. A dip tube allows for sampling at designated intervals.
Offline analysis was conducted using a HPLC, over 35 experimental runs were carried out over a period of 4 months with each experiment being composed of 10 sample points giving over 350 unique data points. The comprehensive HPLC data gave vital information regarding the effect of substrate and enzyme concentration on the process.
The data from this project will allow Oxford Biotrans to further improve their technology, as well as demonstrating that Drochaid Research Services has the capability to successfully support companies in the industrial biotechnology sector.