CAN BIO-SYNTHETIC BIOLOGY BE THE NEXT DISRUPTIVE ... · CAN BIO-SYNTHETIC BIOLOGY BE THE NEXT...

Paper presented at the IFEAT International Conference in Singapore, 4 - 8 November 2012 ‘Essential Asia’ 1

CAN BIO-SYNTHETIC BIOLOGY BE THE NEXT DISRUPTIVE TECHNOLOGY IN TRANSFORMING THE FLAVOUR AND FRAGRANCE LANDSCAPE?

Dr P. M. Murali

Evolva Biotech, Ticel Bio Park Ltd., Taramani Chennai 600 113, Tamil Nadu, India

INTRODUCTION Let me try to make my presentation as exciting as possible, something like a sci-fi, James Bond movie of some sort, so people get absolutely energised about this space. Because I am part of the most frontier science, and there are only about six or eight peers globally. The earlier two speakers have given you quite an insight into the science and cautions so I am going to be talking from a very company perspective on what it is that we have been doing for quite some time now. So if you bear with me for the next 20 minutes, I will try to run through and tell you what is happening in this exciting field. This slide outlines what I am actually planning to cover. To begin with there are going to be two slides about my company, the next one and the last one that will deal with my company. All the other aspects of my talk are about the platform technology that we are using, the type of products that are coming out, and also the benefits of how we can use this science particularly in the area of flavour and fragrances.

So just to give you a perspective of what I am doing and why I am standing here, my company was founded in 2004 and we are an out-and-out venture funded company. We have used about $87 million to date, and the main idea was to develop the platform technology towards some of the applications that I am going to show you today. We have become men from being boys, in the sense that we are now listed on the stock exchange for about four years now with our three letter call sign EVE, which you can easily remember.

Listed in Switzerland we have operations worldwide. We have labs in four places, San Francisco, very close to the Bay area, then Chennai in India, Copenhagen in Denmark and Basel in Switzerland. We employ about 90 people, most of them PhDs, men and women, all enthusiastic and among the younger generation.

Role%of%bio)synthe/c%technology%in%transforming%

the%flavour%&%fragrance%landscape%

P.M.Murali%Managing%Director%

Evolva%Biotech%%%%

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Saffron%

Baker’s%Yeast%

Evolva&Proprietary&Material&

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!  Evolva&introduc3on&

!  Synthe3c&biology&&&bio9synthe3c&technology&

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!  Introduc3on&to&Evolva’s&technology&

!  Key&benefits&

!  Crea3ng&value&through&diverse&but&related&products&

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Slide 1


Introduc)on*to*Company*

!  Founded*in*2004*□  Venture&capital&backed&from&2004&–&2009&

□  Raised&CHF&87m&in&this&period&

!  Listed*on*Swiss*Stock*Exchange*(Main*Standard)**□  Since&December&2009&&

□  Ticker&is&“EVE”&

&

!  Opera)ons*in*Basel,*Bay*Area,*Chennai,*&*Copenhagen*□  c.&95&people&worldwide,&&>70%&with&PhD,&MSc&or&equivalent&

Slide 2

2

Now let me explain the technology, and how we are using it. First of all there are a lot of drivers that have changed this field to a certain extent in recent times. So if you see what the Agilent CEO has been saying, he likens synthetic biology to what semi-conductors did to our lives. A lot of us believe that this is how it is going to be, and the reason is because you can take up a lot of problems in the industry space that can be solved – or can be addressed at least, if not solved – from using some of the technologies that are now currently available. You must have heard from earlier speakers that basically synthetic biology creates artificial biosynthetic pathways. Largely you might have heard that it’s on account of the tremendous advances that sequence technology that has brought the cost down, which means today you can synthesise a gene pretty easily with this information. I do not want to exaggerate that you can do it in the kitchen, but it’s as close as doing it there. Plus there has been a fusion of other types of disciplines that have gone into it, for example, bio-engineering, metabolomics, screening and analytics and informatics. Some of the other areas like fermentation are already pretty industrial. So you see our conviction is the same that others believe, that there is going to be multiple industries that are going to benefit from synthetic biology, namely energy, chemicals, advanced materials, healthcare, nutrition and so on. One of the things my company is doing, just to be very clear here, because you have had slides which talk about costs and its implication. We are where the yellow circles are on the slide: nutrition, consumer health, and pharma. Because we know that is where typically you tend to make more money. So the industry perspective can never be sacrificed when you really want to do some science. Because you can do it in fuels and chemicals, but you know you have to have costs which are going to be downwards of $10/kg, whereas in these spaces you are going to have costs which are going to be $10 upwards. So our company has been focusing on high margin health and wellness sectors as a whole. So that is where my company is actually going. Now I just want to quickly give you an introduction to this, primarily about food and how we see it. So we had these very interesting statistics which said there are possibly about 11,000 basic diverse types of food, and so that would be anywhere from being tea, to oysters, to beer, ginger, truffles, whatever you may call it – I mean, this is how you would be looking at it. And most of us know that food also


Synthe'c)Biology)&)Bio0Synthe'c)Technology)

!  “Synthe(c*biology*..*can*have*as*profound*an*impact*in*the*21st*Century*as*semiconductor[s]*in*the*20th"**□  Bill&Sullivan,&CEO&&Agilent&Tech.&Inc.&(NYSE:&A),&April&2011&&&

!  Creates)ar'ficial)biosynthe'c)pathways))□  Driven&by&extremely&rapid&(Moore’s&law&plus)&&

advances&&in&gene&sequencing&and&synthesis&

□  Plus&a&fusion&of&mulOple&disciplines&P&genome&engineering,&metabolics,&screening,&analyOcs,&informaOcs,&fermentaOon&

&&

!  Will)significantly)change)mul'ple)industries)□  Energy&□  Chemicals&

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□  Healthcare)0)nutri'on,)consumer)health,)pharma))–)Evolva)focus)&

& Slide 4

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Will$Transform$Our$Ability$to$Exploit$the$Cornucopia$of$Food$$

!  Food$is$very$diverse$=$c.$11,000$basic$foods$in$the$world$□  Tea,&oysters,&tomatoes,&guava,&milk,&chickens,&soy&sauce,&honey,&

marmite,&seaweed,&herrings,&&beer,&&ginger,&truffles,&bees&etc.&

!  Food$contains$many$valuable$compounds$□  Some&are&known,&but&many&that&never&been&described&□  Many&food&compounds&have&the&func@onality&of&pharma&libraries&&

!  But$most$of$this$chemistry$is$unreachable$in$pracDce$□  Mass&cul@va@on&may&be&unsuitable&for&any&of&several&reasons&□  Chemical&synthesis&&difficult&,&consumers&resistant&when&feasible&□  Func@on&or&presence&of&molecule&may&be&undiscovered&&

!  SyntheDc$biology$can$unlock$this$diversity$□  And&allow&its&use&in&nutri@on,&consumer&healthcare&and&pharma.&

Slide 5

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Bee$larvae$(Japan)$

Marmite$(England)$

3

contains many valuable compounds, because that’s one of the reasons we really do eat, otherwise why would we do it. So if you see, a lot of these compounds – and if you see this particular sentence here – many food compounds have the functionality of pharma libraries – there are, at least I know the Indians and Chinese have the traditional knowledge here – which they talk about, their food is medicine, medicine is food, so this is an interchangeable way of looking at things. And food obviously has locked up many of these valuable compounds there.

In reality, what happens is, to address the challenge of feeding so many, you can see that most of the chemistry is technically not reachable in practice. Which means you need to have strategies for mass cultivation, sometimes its not possible, you might have to get to chemical synthesis, like you have other types of areas, even this is being resisted by customers. And then you may have a lot of times functions or the presence of molecules that are not yet discovered or not available. So we believe that synthetic biology can unlock a few of this diversity and can be used very effectively in nutrition, consumer healthcare and pharma. So you can see the

illustrations on the side that tell you what exactly is the benefit of using such a science. If you take one of the compounds in the sweetness phase, all of you must have heard and must be using these sweeteners, some of them are artificial, some of them come from natural plants, one of the most popular ones, the next generation of sweeteners, is from Stevia leaves, quite extensively grown in this part of the world and also in China. But it comes with its own little set of, I won’t say drawbacks, but limitations. The problem is that there are certain types of molecules in the Stevia which is sweeter than the rest, but unfortunately they are available in such small qualities that if you are going to be using a conventional method of isolating them through an extraction process or through any other process, you end up not getting enough that is commercially viable. So you may have to then use other methods like this where you can clone all the genes that are responsible, put them in an organism and ferment it. So that’s the type of chase you can do with synthetic biology. The other is the most exotic plant in the universe, namely saffron. So if you have to – and I think some of the Indian cooking today which was left at lunch had those threads of saffron on it, I don’t know how many of you noticed it – but if you see that we need almost a kilo of saffron, and to make a kilo of saffron we need to have 250,000 of these plants picked, so whatever little sweet dish that was there must have been a few thousand plants before you actually get that. And there are other constraints for several countries. For example, 90% of it is grown in Iran, so you might not have a lot of other countries being able to obtain access, so there is a supply chain issue which comes with it. And then you also have issues where the quality has to be maintained, and I know the chief adulterant for saffron threads is a very simple product that comes from the corncob, you know, those threads that are there, they are unadulterated. So you have to solve a lot of issues that are connected in terms of how you make a product.


Will$Transform$Our$Ability$to$Exploit$the$Cornucopia$of$Food$$

!  Food$is$very$diverse$=$c.$11,000$basic$foods$in$the$world$□  Tea,&oysters,&tomatoes,&guava,&milk,&chickens,&soy&sauce,&honey,&

marmite,&seaweed,&herrings,&&beer,&&ginger,&truffles,&bees&etc.&

!  Food$contains$many$$valuable$compounds$□  Some&are&known,but&many&that&never&been&described&□  Many&food&compounds&have&the&func@onality&of&pharma&libraries&&

!  But$most$of$this$chemistry$is$unreachable$in$pracDce$□  Mass&cul@va@on&may&be&unsuitable&for&any&of&several&reasons&□  Chemical&synthesis&&difficult,&consumers&resistant&when&feasible&□  Func@on&or&presence&of&molecule&may&be&undiscovered&&

!  SyntheDc$biology$can$unlock$this$diversity$□  And&allow&its&use&in&nutri@on,&consumer&healthcare&and&pharma.&

Slide 6

$Capsiate$a$promising$weight$loss$agent.$$But$1$part$per$

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Reb$D$<$0.1%$of$stevia$leaf$Very$similar$to$other$molecules$

Very$hard$to$purify$

To$make$1$kg$of$saffron$must$$pick$250,000$crocus$flowers$$

>90%$from$Iran$$

4

So these examples will give you an idea of what sort of compounds and what sort of limitations of these so-called geography and supply chain are limiting its use to commoditise, and those are the things that my company is trying to use. So this is a quick oversimplification of the most complex thing we are doing. Because we started in 2004, and we are still learning every single day, so what do we do? This is nothing but a combinatorial genomics that we assembled in baker’s yeast, so we can identify multiple sources of genes from different types of species, all natural and living. We can stitch them together, put them in yeast, make the desired compound by identifying what we have to do, and then we can de-replicate the process to find out how we have done it. And by this we can make one particular yeast, make the product that we desire, which ultimately can be scaled up. Which means you use beer, I mean every single day a brewery is going to brew tonnes and tonnes of beer there, yeast is the organism, so when you want to make a compound, this is going to be very similar to the way beer is brewed, but instead of making alcohol there you will be making one of the compounds which is required. It’s a very highly IP focused area, and we as a company also spend a lot of time trying to navigate through the maze of patents which are there before you can actually produce something which will be useful for the industry. The next two slides explain how we do it. I have used the example of Rebaudioside This is a problem that was not solved for more than two decades, so for twenty years people have been trying to figure out whether they can clone the genes that are required to make the compound Rebaudioside. We as a company did this in about 18 months using all the resources and the manpower that we have in our hands. The reason we did it is because we knew that it is the most important sweetener molecule in the Stevia family. We felt that they will have to be individually made, because when you put them all together it ends up giving you a bit of an aftertaste which a lot of the beverage companies in the world are not exactly excited about, because their customers do tend to have something in their mouths after they drink their most preferred drink. So one of the ways you have to solve that is to probably separate out the different compounds, and its not easy to do it otherwise, because you have such low quantities, so the best thing to do is to probably synthesize them, put them in yeast and make them ferment. The three bullet points summarise what this technology can do. The first one is that you can build production routes that are not in existence any more. In the sense that if there is an enzyme which has to work faster, it has to have a better conversion, a better industrial proficiency, then you end up doing this through this science.


Evolva&'&Combinatorial&Genomics&in&Baker’s&Yeast&

Slide 7

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5

You can make new types of motorways, which I will tell you in the next slide, and you can build a lot of parts together. If you see here in the animation, that for the same enzyme which can make a compound from a to b, you could have at least about six to 8 of them. So what actually this process does, it chooses the enzyme that is more robust for an industrial application, from a set of genes that makes all these steps. So ultimately you have a yeast that will have something like this: the fourth gene, the fourth one here, the last one here, the third one here, which will make this.


Unique'&'Transforma0ve'Technology'Pla8orm'

!  Can'build'(produc0on)'routes'that'no'one'else'can'□  Combinatorial&gene4cs&to&find&unknown&routes&

□  Unique&enzymes&to&”tunnel&through&mountains”&

!  Can'make'roads'into'motorways'–'all'parts'at'once''□  Titres&of&>&25&grams/litre&for&secondary&metabolites&

!  Can'build'routes'that'lead'to'new'places'□  New&func4ons&H&an4Hmicrobial,&UV&protec4on,&cancer&etc.&

□  New&structures&H&ac4ve&“drug&like”&molecules&in&unique&classes&

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Selected&Other&Products&&Focus(on(becoming(a(specialist(manufacturer(

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Eugenol(2(Cloves(

Curcumin(2(Turmeric(

Anthocyanins(2(Colors(

Resveratrol(2(Red(Wine(

Vanillic(Acid(Benzocaine(2(AnaestheFc(

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Protocatechuic(acid(

Protocat.(Aldehyde((

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6

So what actually it does is to make a more robust microbe that can be used industrially to get yields and titres that are very important. Now, we have achieved titres in certain types of compounds to about 25g plus, which is very very important because if you don’t achieve those yields, commercially this is just academic, you don’t have to do anything but just talk about it. The third and most important point is we can also build routes that take us to something absolutely new. If you don’t see something novel, if you haven’t seen it in nature, you could also build it, in the sense that you could also make compounds that are not present. Now I believe is very important for a few types of functions, like if you wanted to look for an antimicrobial, because a lot of our antibiotics are almost gone, for UV protection, cancer and so on. And the most important thing is after the genome sequencing, you could have multiple targets that needs validation. And you could also end up making drug like molecules that are part of this biology here. The next one is the key. We have built this pathway, just for your information, and we have this pathway being sort of, I mean the products, being evaluated by one of the largest flavour houses, and I’m sure somebody must be sitting here, so that’s already being looked at. And if you see here, basically yeast can make up to this blue, so you see the caption here, so yeast is anyway going to make this compound. So what we are actually telling yeast is, yeast we are going to give you two other genes here, so when you start making glucose, when we push these two genes here, yeast can you make vanillin. And that’s basically what it does. So the yeast now, can, instead of making alcohol, starts making vanillin. Now, once you do that, synthetic biology then opens up something very new. In the sense that once yeast starts making vanillin, the same yeast can be asked to start making other compounds in other areas of application. They do not really have to be in the same place. For example, if this is the major crossroad, and if you are able to put the genes that are responsible for making lets say anthocyanin, which is a colour, you can make the yeast do that.

Or you can make the yeast do something called resveratrol, which is part of the paradox, the French paradox, I mean people are still wondering whether that’s really going to do something about ageing, does it really do something about the way we are living. If you really want to take the necessary dosage of resveratrol that will make you feel happy and young, I am assuming that you will have to drink barrels and barrels of wine. But if you can possibly do this by asking yeast to make this for you, you could have them in a capsule or maybe you could mix it in your favourite drink, as a



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Eugenol(2(Cloves(




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7

small additive, wherein you could have I’m assuming I’m just referring a cola with no specific reference, you could have a cola of resveratrol that’s 45, so you don’t have the guilt of drinking a cola and feel that you’re doing this because it’s a health hazard, or anything like that. So you could make some of these products, and for example you could make a eugenol, you could make a curcumin that is a colour. So any of these possibilities exist once you have yeast cloned, and you can ask it to make a series of other ones. So another example here, you could also be ending up making an anaesthetic, or you could have a vanillic acid coming out, or you could have something from ginger, or something from capsaicin that is chilli that I showed you in the previous slide. Now what makes this more industrially really acceptable is, lets say if you’re going to earn about 600 million dollars in this space, and if you know the first time how to make this. Then incrementally making the same yeast to do a few other types of products is going to keep moving up your bottom line in terms of what you can do. So commercially it makes a lot of sense to use synthetic biology in a way to keep building. So even though these are all having different applications as a whole, you end up having a completely different set of product portfolios that can come out from this. So our idea is to become the focused, specialist manufacturer. And that’s where we have taken this company from where we are.



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Vanilla&Flavors&

Selected&Other&Products&&Focus(on(becoming(a(specialist(manufacturer(

Capsaicin(2(Chilli(

Ethyl(Vanillin((

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Eugenol(2(Cloves(




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8

Now how do we look at it, this is a very familiar picture for a lot of people who have been to England, and who come from there. We have taken a lot of inspiration from the metro picture here. So we look at synthetic biology basically as some sort of metro, so there’s a point-to-point there, and then you could keep extending these points in the biosynthesis by trying to move them all over. From the IP point of view, we would try to control the trunk routes of these pathways, so that people don’t go above and below you, and at the same time you try to see that you are in the position to extend these pathways to make a different set of molecules each time. So it could be in the flavour, in the fragrance, in the wellness, it could be in any space, it could be in perfumery, it could be in a lot of other places. The challenge here is to keep growing this pathway. So we as a company have been growing this.


The$Yeast$Metro$..$enables$many$valuable$products$

9

And you can see that any of these compounds here is technically possible if you know the pathway, and you know how to extend it. And some of the pictures that you have here, we have already done a few of them. So in my last slide I will tell you what those compounds are. So you could make an endless number of compounds that come from here. Which means you’re going to supplement some of the agricultural and supply chain issues which you normally find by using this in a fermentation media.

This will give you an idea of what we are doing. We are actually into biosynthesis and we are focused primarily on nutrition, health and wellness. Fermentation is the way I believe things will go, and this is my opinion. I’m sure others are entitled to theirs. Because of the simple reason that its going to offer the industry a better option of having control over what products they are going to put out into their market. And I believe this is what it’s going to be going into.

As to where our focus is concerned, we are very clear. We are not competing; we are not trying to substitute anything. What we are actually doing is that we are focusing on innovative, high-value and sustainable ingredients. And that is the key to how we are looking at synthetic biology. There could be other people doing other things at this point in time. But we as a company believe that this should be the way in which we should be going, and most of the time I believe that the sustainability is the key to a lot of things, because if you want to really commoditise a few products, particularly say if you


The$Yeast$Metro$

Saffron$

Frankincense$

Mint$

Taxol$

Truffles$Ginseng$Breast$Milk$Carmine$Turmeric$Vanilla$ Resveratrol$Dopamine$

Capsiate$

Caviar$

Ambergris$

Musk$

Stevia$Sandalwood$Cocoa$Opiates$Caffeine$Pyrethrin$

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Evolva&Snapshot&

!  Biosynthesis&company&focused&on&nutri7on,&health&and&wellness&&□  Unique,&proven,&yeast&fermenta7on&technology&□  Strong&differen7a7on&versus&peers.&&>50&patent&families&

!  Focus&on&innova7ve,&high&value,&sustainable&ingredients&□  Four&products&to&reach&market&in&next&few&years&□  Strong&USPs&&F&&cost,&performance,&supply&chain,&etc.&□  Pharma&products&to&be&partnered&&

!  B2B&Model&□  Both&sell&ingredients&and&apply&technology&to&partner&needs&□  Partnerships&with&IFF,&BASF,&RoqueLe,&Roche&□  c.&USD&12&million&of&revenues&in&2011&

!  Many&major&companies&want&to&grow&in&space&□  But&have&limited&partnering&or&acquisi7on&op7ons&&□  Evolva&well&posi7oned&F&differen7ated&assets&and&experienced&team&

Product&Family& On&Market&&Addressable&Market&&

Vanilla& 2013& &$&0.6&bn&&&

Pomecin& 2014& $&1&bn&

Stevia& 2015& $&4&bn&

Saffron& 2015& $&1N2&bn&

10

want to do saffron, availability cannot be a limiting factor in terms of how it can be put to a lot of human beings in the world. So this is one of the things that we are very very keen on. You can see that is one of the reasons we chased the four types of products listed. This is what is likely to be in the market between 2013 to 2015, so we are not probably in the next generation, or we are not in 2030, because these are all compounds which we believe will take us very quickly there. Of course there are all these regulatory issues that I do not want to get into, which we are presently doing, and pursuing and things like that. Vanillin because we believe that the natural vanillin is so expensive, the synthetic version is certain not a very preferred choice for a lot of people. So this is giving us a very niche space wherein we are giving them a fermented vanillin. We have done a taste panel, we have evaluated it, some of the leading players have checked it and they find that this is so good that it is as good as what they have seen possibly in the natural space. Then we have a set of anti-fungals, which are absolutely an inspiration from pomegranate, and you know this is now becoming another plant that is so good. And that’s another product, and we have some anti-fungals that we are using and want to use for the food industry primarily. Stevia, as you know, is going to be the next big thing which is happening in the world and we are very much there. We have made Rebaudioside D in sample forms, and it is been tested by a lot of people, particularly some of the companies that are sitting here, and they feel this is absolutely good. I think we are the first in the world to have a very high purity Rebaudioside D, even for testing purposes or evaluation purposes by a panel of people who could actually taste it. So this is one of the other products, so this is likely to come out after all its regulatory issues in 2015. Saffron, obviously that is something that I really think has been a game changer in terms of how we have used the synthetic biology. There are three compounds which constitute what is within what is called saffron, the smell, the colour and the taste and we have separated out all the three, picked out the genes which are responsible, and put it into yeast. So technically this yeast now can make all the three separately, it can make the colour, it can make the aroma, it can make the taste as well. I mean its absolutely amazing because you really don’t need to go and get 250,000 plants to get a kilo, and you really don’t have to be in Iran to harvest any of this. So this is one of the things that you may like to know. We have a very interesting model. We are a B2B company, which means obviously we do not go out to the people there, we sell our ingredients and we apply our technology. Among the partnerships I think the previous speaker mentioned BASF so I assume they have moved to Switzerland now, more than to Germany, so they are working with us on this, and this is hardcore synthetic biology again. So we presume that, as a company of about 8 years old, we are actually addressing some of the unmet needs in the industry, and we are chasing compounds which will be really useful and which can be industrially scaled up, so this explains to you in a nutshell what we are doing. And just to give you an indication, that these compounds through fermentation is not really far off, I mean they are just about a couple of years away. In the USA, I believe there are already a few compounds which are through the same technology, and so I think you are going to see more of these companies coming in in the future, in terms of how they can assembly pathways and put them together to get to the stage where they can industrialise them and make them available through fermentation routes for companies. So this is all I wanted to present today, thank you very much for your attention.

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Dr. P M Murali, is the Managing Director of the Swiss International Discovery Company Evolva Biotech. He established Evolva Biotech as India's unique public private partnership in Hyderabad, India before the facility moved to Chennai. Dr. Murali has a Ph.D. in Microbiology from Madurai Kamaraj University and a D.Sc. with over 20 years of experience in Pharmaceutical/FMCG R&D. He was the Founder and Director of the Dalmia Centre for Research and Development for 16 years and prior to this he has worked as an Indo-US researcher at the Battelle-Kettering Research Centre, Ohio, USA. He has a long track record in launching innovative products in the

FMCG space. He serves as the Chairman of the Biotech panel for Tamil Nadu with the Confederation of Indian Industries Tamil Nadu and also as the President of Association of Biotech Led Enterprises (ABLE) the only industry body for biotechnology in India. Dr. Murali guides students for their Ph.D. at the Bharathiar University and is an Adjunct Professor of the Deakin University in Australia.

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