Industrial Saponins

Lead Research Organisation: University of Edinburgh
Department Name: Sch of Biological Sciences

Abstract

The aim of this project is to bring saponin production into the domain of industrial biotechnology to make this diverse group
of biosurfactants available for commercial exploitation at the required scale, structure specificity & cost, in the first instance
for home & personal care (HPC) use. We have already demonstrated that saponins can synergise with conventional
surfactants to reduce the total surfactant level required for cleaning performance in laundry detergents, and that such
mixtures are particularly effective at low temperatures. Saponins are plant-derived triterpenoidal or steroidal surfactants.
Saponins currently have limited commercial availability, being sourced from natural plant materials. This supply is deficient
in scale, is of limited structure specificity and is too expensive for Unilever's intended use in laundry detergents. Unilever's
concentrated detergents business alone would require 100 tonnes pa of saponin at 0.1% level and 20% product range
penetration. Leading saponin supplier Desert King produces only 20 tonnes active saponin pa from natural extracts.
Furthermore, we have demonstrated that certain saponin structures are significantly more effective than others. The current
sources of saponins are natural extracts and comprise a heterogeneous mixture of saponin structures. Our IB innovation
will enable considerable enrichment for the most effective saponin structures. Current commercial saponins are too
expensive at $35/kg for crude extracts for use in anything other than premium products. We expect IB to be a more cost
effective route to saponin supply.

Technical Summary

In a current project (TSB 131168), we have demonstrated production of tailored saponins in Nicotiana benthamiana and
have expanded the synthetic biology tool kit genes and enzymes for tailored saponin bioengineering. However, this
tobacco platform is not sufficiently scalable for commercial production. We need to transfer the saponin synthetic pathways
into a microbial platform. We have selected yeast, Saccharomyces cerevisiae, as the most appropriate platform. Our aims are to push science and technology boundaries in multiple dimensions: (1) to transfer saponin metabolic pathways from
model plant platform to yeast and engineer the pathway for commercially viable yield; although an academic proof of
concept of simple monoglycosylated saponin production in yeast exists, further innovation is required to enable complex
hyperglycosylated saponin production and to tackle yield & flux optimisation; (2) for downstream process development and
scale up; (3) to further develop the triterpene toolbox of enzymes, genes and pathways for specific saponin structure
enrichment & production; (4) to further understand the solution microstructure and surface adsorbed layer properties of
mixtures of saponin & conventional surfactants, and (5) to continue to develop tangible benefits and structure-function
relationships demonstrated in TSB 131168 for inclusion of saponins in model HPC formulations, extending the complexity
of the models to include associated detergent technologies such as enzymes & dispersants. The above innovations will
collectively enable a powerful business case for saponin commercialisation to be constructed.

Planned Impact

As described in proposal submitted to IUK.
 
Title Image of Nicotiana benthamiana 
Description Nicotiana benthamiana, a relative of the tobacco plant, is commonly used in plant research. Here, the leaf is used as a host organism to produce components of avenacin, a fluorescent antimicrobial produced by oat roots, which protects the roots from soil pathogens. A cluster of genes - the Sad genes - have been identified in oat as the instructions used to produce avenacin. The blue spots indicate the presence of parts of the avenacin compound introduced by inserting some of the Sad genes into the host plant. The other spots are controls to test the expression system. Image supplied by Aymeric Leveau, Osbourn laboratory. 
Type Of Art Image 
Year Produced 2015 
Impact Image placed on the Norwich Research Park Image Library, for free sharing and dissemination. 
URL http://images.norwichresearchpark.ac.uk/imagedetails.aspx?imgid=103
 
Title Winner of Norwich Research Park Image Library Competition: Dr Thomas Louveau 
Description More than 150 images of work were submitted by researchers, clinicians and scientists who work at the site, for the NRP image library competition. These were narrowed down to 12 finalists - which will form part of a calendar - from which an overall winner was selected. The winning image, entitled A New Planet, was taken by Thomas Louveau, a post-doctoral scientist in the Department of Metabolic Biology at the John Innes Centre. The image show a dried plant extract in a round flask taken from behind on a dark background. The extraction is part of the purification process of new-to-nature metabolites produced by synthetic biology approaches. The new metabolic pathway has been engineered in Nicotiana benthamiana, a relative of tobacco by scientists at the John Innes Centre on the Norwich Research Park who are studying natural products.. 
Type Of Art Image 
Year Produced 2015 
Impact A calendar was produced from the 12 final images, with "A New Planet" on the cover. The images are also displayed in the Centrum building on the Norwich Research Park. 
URL http://edp24.co.uk/news/tech/sheer_beauty_of_science_showcased_in_competition_at_norwich_research_pa...
 
Description A plant derived triterpene toolkit of biosynthetic genes has been identified, cloned and tested for activity in Nicotiana benthamiana. A combinatorial approach has been taken and a range of saponins with diverse structures and properties have been shown to be produced.

We have engineered saponin biosynthetic pathway genes from a variety of plants for expression in yeast hosts. We have demonstrated the expression of these plant derived genes and saponin production by the engineered yeast.
Exploitation Route The project is in collaboration with Unilever who are looking at the potential uses of saponins in their home and personal care business.
Sectors Chemicals,Environment,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

 
Description The project's industrial partner has filed a number of related patents. One of the academic collaborators is in the process of investigating that feasibility of spinning out a company. Further academic-industrial collaborative project funding has been obtained
First Year Of Impact 2019
Sector Agriculture, Food and Drink,Chemicals,Manufacturing, including Industrial Biotechology
Impact Types Economic

 
Description Bio-engineering non-sugar modifications of saponins - BBSRC CTP studentship
Amount £41,000 (GBP)
Funding ID BB/R505584/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 11/2017 
End 10/2021
 
Description Engineering yeast organelles for production of saponins
Amount £63,600 (GBP)
Organisation Industrial Biotechnology Innovation Centre 
Sector Academic/University
Country United Kingdom
Start 10/2019 
End 09/2022
 
Description Harnessing enzymes from plants for selective functionalisation of triterpenoid scaffolds
Amount £386,779 (GBP)
Funding ID BB/S017712/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 07/2019 
End 06/2022
 
Description Industrial Biotechnology Innovation Centre - SynBio Accelerator project
Amount £200,393 (GBP)
Organisation IBioIC 
Sector Academic/University
Country United Kingdom
Start 06/2017 
End 05/2019
 
Title Transient plant expression technology for triterpene production at preparative scale 
Description We have develop a method for transient heterologous expression of biosynthetic enzymes in N. benthamiana for production of high-value triterpenes. Agro-infiltration is an efficient and simple means of achieving transient expression in N. benthamiana. The process involves infiltration of plant leaves with a suspension of Agrobacterium tumefaciens carrying the expression construct(s) of interest. Co-infiltration of an additional A. tumefaciens strain carrying an expression construct encoding an enzyme that boosts precursor supply significantly increases yields. After a period of five days, the infiltrated leaf material can be harvested and processed to extract and isolate the resulting triterpene product(s). This is a process that is linearly and reliably scalable, simply by increasing the number of plants used in the experiment. We have developed a protocol for rapid preparative-scale production of triterpenes utilizing this plant-based platform. The protocol utilizes an easily replicable vacuum infiltration apparatus, which allows the simultaneous infiltration of up to four plants, enabling batch-wise infiltration of hundreds of plants in a short period of time. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? Yes  
Impact Generation of gram-scale quantities of 98% pure triterpenes and demonstration that we can rapidly carry out combinatorial expression of enzymes from our triterpene toolkit to generate known and new-to-nature compounds. This had attracted considerable interest from industry and led to four new projects directly funded by different companies in the pharma, ag, food and drink and home and personal care sectors. 
URL https://www.jove.com/video/58169/transient-expression-nicotiana-benthamiana-leaves-for-triterpene
 
Description Built new image library for the Norwich Research Park 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact To help raise awareness of the high quality research carried out across the Norwich Research Park, we developed an open access image library to showcase images of NRP science accompanied by accessible legends suitable for a general audience. The image library has the potential to attract new traffic to the NRP institute websites and research group homepages and will allow groups such as the media, schools, the general public and other researchers to gain a glimpse of the great depth and diversity of research that is being carried out in Norwich. Although all the images are freely available to download and use, people are required to sign up to use the library and are asked to enter information at the time of download about their intended use of the image. This helps to capture who is using the library and where the images are being used. So far images have been downloaded 582 times and have been used in presentations, publications, on websites, for education and advertising. We ran an image competition to raise awareness across the NRP site and then assembled a judging panel to select the top 12 images which were used to create an NRP calendar for 2016. The calendar was sent to politicians, business leaders, industry and academics at institutions in the UK and internationally. The competition featured in the local paper, the Eastern Daily Press and an overall winner was picked and used to make a large canvas which is hanging in the new Centrum building. We have received many positive comments about the libraries ease of use for finding good quality, copyright free images to use in presentations from fellow researchers across the site. In October 2016 we put on a two-week, large scale image exhibition at the Forum in Norwich as part of the first Norwich Science Festival. The images attracted a lot of interest and several artists have been in touch with scientists whose research images were on display to set up new collaborations using science to inform artistic practice. We will post outcomes of these endeavours onto the image library website to widen our exposure to new audiences.
Year(s) Of Engagement Activity 2015,2016
URL http://images.norwichresearchpark.ac.uk/
 
Description Great British Bioscience Festival 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact We were successful in a winning a BBSRC science communication award to develop a new science stand called 'Nature's Factories' to run at local events and as part of the BBSRC's Great British Bioscience Festival showcasing the best of British bioscience in its 20th anniversary year.
Human evolution is tightly linked to our use of plants for food, building materials, fuel, medicines etc., and we continue to look for innovative ways to use natural resources to provide us with sustainable solutions that support our lifestyles. Scientific research is enabling us to discover and develop new plant products that improve our lifestyles by creating better medicines, healthier foods and greener technologies whilst also demonstrating the importance of protecting plant species diversity and ecosystems.
The Nature's Factories stand was designed to enable the public to find out how science is exploring and exploiting the valuable variety of chemicals made by plants as well as making natural remedies to take away and pick leaves from our fact-tree.

We took the stand to the Science in Norwich Day at the Forum in Norwich (1st June 2014), to the Cambridge Botanic Gardens (19th August 2014) and on the 12th November 2014 used the stand for a training workshop for EU scientists to learn how to create interactive displays for public events. On 14-16 November 2014, our exhibit headed to Museum Gardens, Bethnal Green, London to be showcased at BBSRC's Great British Bioscience Festival. The festival, delivered in partnership with London Science Festival was open to the general public and free to attend.
Since then the exhibit has featured at the Bury St Edmunds family science festival (21st March 2015), the Fascination of Plants day at the John Innes Centre (14th May 2015), the Festival of Plants at Cambridge Botanic Gardens (16th May 2015) and at the Youth Stem Awards (13th January 2016). We plan to use the exhibit throughout 2016.
Year(s) Of Engagement Activity 2014,2015,2016
URL http://www.bbsrc.ac.uk/engagement/exhibitions/gb-bioscience-festival/
 
Description High Value Chemicals from Plants- Harnessing the potential of synthetic biology for industrial biotechnology. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact High Value Chemicals from Plants- Harnessing the potential of synthetic biology for industrial biotechnology. 13-14 July 2015, Dunston Hall, Norwich. I delivered a 20min talk about the use of the HyperTrans system in goldengate vectors and its benefits for metabolic engineering.
Year(s) Of Engagement Activity 2015
 
Description NRP DTP Summer Conference 2015 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Professional Practitioners
Results and Impact NRP DTP Summer Conference 2015 held in the assembly house in Norwich. 15 min talk: Pathway engineering using the GoldenGate MoClo system.

Norwich Research Park Doctoral Training Partnerships
Year(s) Of Engagement Activity 2015
URL http://www.biodtp.norwichresearchpark.ac.uk/
 
Description Norwich Science Festival John Innes Centre Exhibit 22.10.16 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Norwich Science Festival.

The interactive 'Power of Plants' stand from the Latitude Festival made another appearance at the first Norwich Science Festival with Katrin Geisler coming over from Cambridge University to join JIC scientists Michael Stephenson, Hannah Griffiths, Colette Matthewman, Zhenhua Liu, Nadiatul Radzman, Dorota Jakubczyk, Don Nguyen, Miriam Walden, Jenni Rant and Roger Castells-Graells for two days as part of the Norwich Research Park's 'solving problems with science' weekend. A new addition to the stand was developed by Roger who introduced visitors to the structure of viruses through his challenge to build a giant virus particle. Plants are being used to produce virus particles for use as vaccines and so for visitors this deeper look at viruses in combination with a hands-on experience of mock infiltration of Nicotiana benthamiana plants with water enabled the scientists to explain the techniques involved and exactly what is being made by plants in this system. People were very interested in the variety of science on display and asked lots of questions. It was good to showcase our research at the first Norwich Science Festival and to show people locally how far reaching the work taking place in essentially a rural county can be in terms of impact to societies across the globe.
Year(s) Of Engagement Activity 2016
URL http://norwichsciencefestival.co.uk/
 
Description Science in Norwich day 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Helped Jenni Rant (SAW) to hold a stand describing the scientific research ongoing at JIC, at the Science in Norwich Day at the Forum on the 1st of June 2014 .
Year(s) Of Engagement Activity 2014
URL http://soin.org.uk/2014/04/science-in-norwich-day-2014/
 
Description Science, Art and Writing (SAW) project on plant natural products, Suffolk 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact A Science, Art and Writing (SAW) project on plant natural products was run for 90 students at St Benedicts Secondary School, Suffolk, UK on 3 July 2015. The students collected plant samples from the school grounds and used them in chromatography experiments to see how many different types of pigments they could extract.
Year(s) Of Engagement Activity 2015
 
Description Submitted scientific images to the Norwich Research Park image library 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact The NRP image library consists of images arising from research in life and environmental sciences on the Norwich Research Park . The aim of NRP research is to deliver solutions to the global challenges of healthy ageing, food and energy security, sustainability and environmental change and this is reflected in the images in the library.
Year(s) Of Engagement Activity 2016
URL http://images.norwichresearchpark.ac.uk/searchresults.aspx?query=leveau