Full transcript

[00:00:00.000]
I’m going to talk about a project that I worked on a few years ago. It’s all old news in a certain way, but it will be new to you. It’s called a Biot Prospecting for Psychotherapeutic Drugs in the Amazon Rainforest. As you can see there, I’m going to sit down most of the time, I guess. Mental and emotional disorders takes a huge toll on society. These figures are out of date, but you get the idea. There are 37 million physician office visits in 1997 for mental disorders, many for depression, anxiety, schizophrenia, alcoholism, drug abuse. All of these things are enormous health problems on a global scale and take a great deal of resources and are just really a terrible thing. The incidences of these things are going up. Well, you think there would be a great deal of effort to discover new and better medicines to treat these kinds of things, but the fact is there have been no real significant breakthroughs in psychotropic drug development for about the last 30 years. There are plenty of psychopharmaceuticals out there, but many of them are just the same thing in a different package. Companies will tweak the molecules and bring out a new medicine which is really not different than the old medicines.

[00:01:50.090]
But they can call it a different name and they can sell it under that. Not a lot of innovation is going on when it comes to psychotherapeutic drug discovery. Whatever you may feel about the potential of drugs to cure these kinds of problems. But if we turn to ethno pharmacology, there is potentially a rich area there to explore, to look for new psychotherapeutic medicines. And as I said in my first talk, I said, Why should we care? Why should we bother discovery of how psychoactive plants mediate their actions in humans? And I think Dr. Molina said it right. As I said previously, novel chemistries from the natural world leads often to new lead structures, and as often as not, they lead to new pharmacologies or new understanding of pharmacological mechanisms. So it’s not that the natural products themselves may end up in the clinics, but what you learn from them as they’re used in traditional context may lead to the discovery of new medicines that do have clinical efficacy. Dr. Mellina referred to this ancient and horri paper I wrote in 1996, Plant Hallucinogenes, Spring Mortgage for Psychotherapeutic Drug Discovery. I think that was for the Serotonin Conference.

[00:03:18.060]
If it wasn’t for the kind intercession of Dr. Nichols and Mark Geier, this thing probably never would have seen print. Maybe it shouldn’t have seen print, but anyway, I got it printed. Investigating plants, potentially novel psychoactive plants, validates molecular targets for drug discovery. Psychoactive plants are by definition, psychoactive. There’s a wealth of anecdotal data from traditional culture that’s available. In essence, there’s been an uncontrolled experiment going on for thousands of years. That knowledge has been accumulated, although not in any systematic way. But psycho activity is a more rewarding target for drug discovery, this ethno botany driven drug discovery than other types of activity, such as anti cancer activity, for example. If someone tells you that a plant is used to cure cancer, you can’t really tell until you’ve taken it into the laboratory and run it through the appropriate assays if they’re pulling your leg or not. If someone tells you that this plant relieves pain or causes vision or relieves anxiety or whatever, you can test that in the field as long as you’re not too reckless and confirm that they’re giving you the right information or not. There are compelling reasons exist to investigate natural products for psychotherapeutic activity.

[00:05:06.410]
For one thing, 80 % of the world’s population relies on traditional medicine, still for basic health care needs. Folk medicine, traditional medicine is really quite important on a global scale. Mental health is something that happens in every society and every culture, and it’s recognized. Almost every culture has remedies for this. They rely on plants to ameliorate these problems. There’s this vast ethnographic literature on essentially clinically relevant activities, and the psycho activity of the plants is easily ascertained in the field. This was something that struck me about Dr. Mellina’s talk was the categories that we in biomedicine might apply to mental illness may differ from culturally recognized categories. I wanted to follow that up. What we call an antidepressant, there may be cultures where depression per se is not even recognized, but we’re not going to talk about that today. Another good reason is that these ecosystems are threatened. All this molecular biodiversity and botanical biodiversity that are the raw material for exploration in any search for drugs is diminishing, and it’s diminishing rapidly. You’re all familiar with this. It’s important to try to sort through these biomes before these species disappear off the face of the planet.

[00:06:51.830]
That’s happening every day and it’s accelerating. There’s really no way to put a dollar value on what is being lost in terms of its value every day, but it’s enormous. And if we knew how much it was, we’d probably need one of those antidepressants because it is a very sad situation you know, why we’re not preserving our ethno pharmacological treasure house, if you will. So when we turn to the Amazon, the Amazon, for whatever reason, it’s mostly cultural, is a very rich place if you want to look for psychoactive species. Amazonian shamanism and the traditions that are practiced there means that they’re familiar with a great pharmacopia of psychoactive plants. There are other places you can look in the world, certainly, but the Amazon is a rich one. The shamanists themselves are, in a way, experimental psychopharmacologists. They approach their material medica in a very in a very experimental way. They try it themselves. They try it on their patients. It’s not a systematic approach by any means. It’s more like trial and error. But discoveries are made this way. And another problem is it’s not often preserved, it’s not often preserved. It’s not written down. These are oral traditions and so on.

[00:08:20.710]
But it’s not unreasonable to suppose that the knowledge accumulated in these curative traditions, these shamanic traditions, include remedies for cognitive dysfunction, other types of mental problems. A lot of the way you have to go about this is you have to spend time in the field. You have to send good anthropologists, good ethnographers, and ethnobotanists out to the field with the patience to sit and listen to these informants, as Dr. Lune has done so much with Don Emilio Andrade and the others. You just have to be willing to spend time, hang out with these people, and learn from them. That information gathering, but then once you get the information, what do you do with that? How can you take that to the next level? If we’re looking for a cure for schizophrenia, for example, or a cure for dementia, or a cure for depression, or all of these things that our global culture is plagued with, how do you take the next step once you’ve got the ethno pharmacological data? Well, the approach that I took, my colleagues and I took, I should say, in this approach was to look at the receptor role. I was lucky a few years ago to get a grant from the Stanley Medical Research Institute, which is a nonprofit organization that supports schizophrenia research.

[00:10:01.890]
I went to a conference that they gave at the National Institute on Drug Abuse with the unlikely title of potentially therapeutic applications for illicit psychoactive plants. It was something like that. Very strange title. So I went and I gave my usual shtick about Ayahuasca. And then after the conference, I was contacted by one of the people from the granting agency. He said, I liked your talk and we’re interested in natural products. We’re looking for better medicines. We have a project going on in China right now to sort through the Chinese pharmacopeia, but we have nothing going in South America. Are you potentially interested? I said, Well, sure, tell me more. He said, Well, we did fund $136 million in this research in the last five years. Basically, I said, Where do I sign? It wasn’t quite like that, but I said, Okay, you’re in five need me to apply for a grant. That doesn’t usually happen. So I did apply for a grant, and lo and behold, we got it. I enlisted some very accomplished colleagues in this, Dr. Brian Roth, primarily at the University of North Carolina at Chapel Hill. Now, where Dave is currently enjoying his retirement, I guess, he doesn’t talk much about what he’s doing there, but he has an affili with Brian, and Brian is a well known serotonin pharmacologist, and he also runs the National Institute of Mental Health Psychoactive Drug Screening Program, which is one of the few instances where your tax dollars are probably doing some good.

[00:12:03.870]
So I knew Brian and I knew Dr. Roth, and I said, Maybe we could approach this through receptor screening. And he’s talked about the receptor ome, which he’s the world’s expert on. The receptor ome being the entire complement of receptors in the genome, including the non G protein coupled receptors and transporters. Most of the receptors of interest for this project are G protein coupled receptors. But the goal was to screen the receptor room or some significant fraction of it to discover how psychoactive plants might induce their actions in humans and possibly find some new compounds of interest. I’d like to thank Dr. Roth for the following few slides. This is one way you can depict the human genome. These pie shape slices are assigned to different gene complexes and different functionalities, essentially. The blue slice here is the part of the genome that’s devoted to signal transduction. That would be neurotransmission, hormone target interactions. Any signal transducing molecules would come somewhere in this. The receptors represent about one 20th of the genome. And of that fraction, the G protein coupled receptors, which are the primary targets of this project, is only about 2 % of the genome transporters, such as the serotonin transporters, only about 1.7 % of the genome.

[00:13:51.920]
And if you then focus in even more in more detail, the so called group A, G protein coupled receptors comprise a family of about 274 receptors, and the biogenic amine receptors are just a small fraction of that. And this small fraction of what you might call the biogenic amine receptors, the serotonin receptors, dopamine, acetylcholine, and so on, are a small part of this signal transduction complex of genes. Those are the ones that we focused on primarily. We used a technique that may be familiar to I know will be familiar to a lot of you so called radio ligand receptor binding. This is a widely applied technique and as they say, the devil’s in the details. But overall, it’s a very simple thing. If you have a radiolabelled ligand, the radiolabelled drug, essentially, that usually is labeled with tritium. You can incubate it in vitro with brain tissue, or these days they use cell lines with that express specific receptors on the surface of the cell. They only have one type of receptor. You simply try to add. You add increasing concentrations of unlabeled drug. You have your labeled drug up here, in this case, titanic acid, which is an oxytocin.

[00:15:36.130]
You add another compound and you determine the level that inhibits half the binding. By various calculations, you can determine essentially the affinity of the drug for that receptor. Affinity is what you want. You want low affinities. Micromolar, nanomolar or lower. That’s what you’re looking for when you do this broad screening, and you have to decide what’s the cutoff. You look for the high affinity compounds. In Dr. Ross’ laboratory, they screen routinely for many, many of these neurotransmitters. The serotonin receptors, the dopamine, muscarinic, histamine, the transporters, the opiate receptors, all of these, we didn’t screen for every receptor on this list. I think when it was made, they now have many more screens, but we screened for a good swath of them. We looked at the receptors that are known to be associated with some of these mental disorders. We had quite a palette. Essentially, what we’re doing in the study is we’re bringing together the ability to look broadly at these extracts, and then we’re bringing the molecular diversity that you find in the Amazon together. And we’re trying to bring these huge data sets, essentially, together and see if anything emerges out of it that might be worthy of further pursuit.

[00:17:25.610]
And eventually, isolation and development of a new medicine. Another important resource that we had access to at the time was the NAPR Alert database that is a natural products database that’s maintained at the College of Pharmacy at the University of Chicago. And it’s fallen into disrepair in the years that have passed. But it was very extremely useful because NAP alert contains information basically on the traditional uses of plants, the chemistry, and the pharmacology of plants and plant extracts. So it’s a good place to start. They have many different activity codes. They have over 600 different activity codes, these five digit codes. And so we just selected out the ones that have to do with CNS activity, broad range. So the 11s here are the first two digits are all 11, you noticed. So that defines the activity. And then these others are specific for different type. So first of all, we went through the NAP alert database, and using these activity codes, we pulled out the genera from the Amazon that might be interesting based on their activity. You can also get information about the chemistry, the predominant compounds in these genera if they have been looked at.

[00:19:02.080]
And of course, if you’re looking at the Amazonian biome, there are huge gaps. A lot of this has not been well investigated. There’s a great deal of guesswork that goes on with this, but you have to start somewhere. This is just an examination or a visual diagram of how we do it. This is actually for the abstract of the paper that we eventually published in the Journal of Ethical Pharmacology. They now like you to give a visual abstract. It’s pretty simple. It really should begin here over at the project database because we did a lot of the We did the literature search and data collection first in a way. We had an idea what to look for before we went to the field. Then we did go to the field. Most of this work was done around Iquitos. My colleague Juan Ruiz, who is the director of the herbarium there, was really the one that made it possible. We went to the field, we made collections and also herbarium voucher specimens of everything we collected, the quintessential and absolutely necessary herbarium specimen, which were deposited in several institutions. Then once we had identifications, we went back to NAPR Alert and we ran what they call those three part profiles, chemistry, pharmacology, folk use, and collected those files.

[00:20:41.670]
At the same time, we were doing the literature, we were also working out an extraction protocol, most of which was done in the laboratory at the University of Minnesota. My other co PI on this project was Dr. Thomas Hoey, who is a natural products chemist in the chemistry department at the University of Minnesota. So you make the crude extracts, you fractionate them, maybe or not. But anyway, you get them into one of these assay plates. And you saw, if you watched the film last night, Dr. Ross Lamb has these automated roboticized screening technologies set up so they can run many, many extracts through in a very short time. So high throughput screening, you get the data, you analyze the data, you go back to the database, try and validate that. Eventually, hopefully, at the end of all this, you end up with a publication, at least, and maybe some further directions. Let me sit down. My chair is very squeaky. Oh, yes. We have more squeaky chairs than not squeaky chairs. That was the approach. I’m not going to show you much data here because for one thing, it’s hard to represent it there. What you end up is huge tables, and I’m not going to show you slide after slide.

[00:22:18.080]
But just a couple of this might be interesting. One of the things, again, from the NAP alert data was the folk use profile of the plants we collected. We casted a wide net in terms of what might constitute CMS activity or psycho activity. Here we’ve got some obvious things. This is really not a very good pointer, like analgesic, anxiolytic, stimulant, about 30 categories or so. Some of them not so obvious, like magic plants. I can’t see it here and I can’t point it out because this thing is next to useless. Then we also had data on the occurrence of secondary compounds in these collections that we’d made. I should say in the previous slide, I talked about collected species and targeted species. Targeted is basically our wish list from the database, what we wanted to acquire. Collected is what we actually got in the field, so obviously they’re not the same. A lot of times we collected species that weren’t necessarily on our list, and a lot of species that were on our list, we did not obtain. Then besides the folk uses, we also looked at the relative occurrence of secondary compounds. Again, as reported from the literature, alkaloids of any type, diterpines, indole alkaloids, different subcategories of secondary compounds.

[00:24:14.540]
That would give us an idea of what the active principles might be. But this is all based on previously recorded data. We didn’t do all that fractionation. We work mostly, except in a few cases, we work mostly with crude extracts or semi purified extracts. This is the key slide. If you want to look at this work from the 30,000 foot level. You’ve got receptors that were assayed on the vertical axis here, all the serotonin receptors, 1A, 1B, and so on. The alpha receptors, dopamine, the opiate receptors, the muscarinic receptors, the transporters, dopamine, the norepinephrine, and serotonin transporters. Then we just basically looked at it in a fairly descriptive manner. The legend is the % of the red is the % of the total fractions that we collected. The blue is the % of the hit fractions. The hit fractions meaning that they bound to the receptor at a fairly high affinity. They were, I guess you could say, below the cut off. They were considered hits if there was a certain degree of inhibition. Interesting patterns here. I don’t know. Again, this was hardly an unbiased sample, but the 5HT receptors, any 5HT receptor at the very top had 80 % of the hits in our assays hit one or more of the serotonin receptors.

[00:26:17.800]
5ht activity is apparently quite common in this particular sample. I don’t know if that reflects what’s going on in the actual biome because certainly we were selecting for plants that were likely to have this activity. But there are some other surprises here. For example, the opiate receptors, the three opiates here, mu, delta. Well, actually, I guess we just looked at mu and delta, but a surprising number of our extracts seem to hit one or both of those receptors, which I thought was interesting. The transporters, we looked at the different transporters and didn’t get a lot of activity, but it was interesting that it was there. Maybe there’s a lot of opiates out there. Then on a fairly small fraction of these collections, we did what was called functional assays. The binding assays don’t tell you if a drug is an agonist or an antagonist, really. It just tells you that it binds. You have to do a further functional assay to essentially answer the question, when it binds, does it have an action? Does something happen? Does it elicit a second messenger, or does it just block the receptor? So we did functional assays on a few of the collections that we had that had the highest 5HT2A affinity in the binding sites.

[00:28:03.000]
The reason, you all know probably that we selected 5H2A is because that’s one of the primary receptors for hallucinogenes. This grant was really not to discover new hallucinogenes. Well, that may have been the subtext of the project, but the rationale of the project was to find new compounds that could address the so called negative symptoms of schizophrenia. Not overt psychosis, but all the other symptoms associated with schizophrenia that cognitive deficits mostly. Psychoph clinics may not be psychotic if they’re properly medicated, but they may not be particularly functional either because they have cognitive and memory problems. That neatly grades over into dementia, which is a burgeoning problem. So essentially get that as a target as well. We screened some plants, and we screened all the plants at the 5HT2A receptors. Then the ones that had the highest affinity, we screened in these functional assays, thinking that if they were agonists in these assays, then they might be potentially hallucinogenic. In fact, many of them had been reported to be hallucinogenic in the ethno botanical literature, but hadn’t really been demonstrated. There was not a bad match between the folk literature and the probable hallucinogenic activity based on these binding activities.

[00:29:59.040]
Someone yesterday say, Oh, yeah. I think in the film last night, Sarah Hart was interviewing Dr. Roth, and he was saying, Well, it turns out that the 5HT2A receptor is a primary target for the so called second generation antipsychotics. They have different receptor profiles, but the 5HT2A receptor is a primary target for the so called second generation antipsychotics. Dr. Roth was saying, Well, out of this screening might emerge a compound or more. If 5HT2A activation is a hallucinogenic effect, maybe some of these compounds will block that and then potentially be useful for mental health. But I had to think back when I started my postdoc at NIH many years ago, we were working on 5HT2A receptors, and my supervisor was saying, Yes, and maybe out of this we’ll find a compound that is an anti hallucinogen that we can use to block the hallucinogenic effect. I was very disappointed. I said, Dr. Sevedra, I thought we were looking for better hallucinogen. He was not amused. These genera emerged as interesting. Then there was actually, ironically, an interesting, Schultes wrote in the early 90s about plants that he had collected over his years in the Amazon that could potentially be used to treat dementias because he was coming down with dementia.

[00:31:51.170]
Rather a sad story. He knew that he was losing it in a certain way. In some ways, this was his gesture to that and his recognition of it. He wrote about these plants, particularly, that had been reported to be used for dementia. We screened those with particular interest, and no particular patterns emerged, except the taverne montanas appeared to have the right profile. The taverne montana is a very close relative of Eboga. It’s in the Pocinacea. This genus is really well recognized for complex indole alkaloids. But this isn’t really a discovery because it’s known that these things have these alkaloids and they often work on the ser atonal receptors. But in terms of identifying a genus to focus on, if you wanted to do further isolation and bio assay and behavioral pharmacology or whatever, the tavern, they Montana’s Emerge is definitely promising. There were about six others that could be followed up, although our grant never got that far. That’s a quick thumbnail look at the science, what we got out of this. I hope that they were happy with it. We did eventually publish, but I’ll give you an idea of how it goes in the field.

[00:33:33.500]
This is Iquitos, this is Belem, which is the community in Iquitos that’s close to the river. This is where most of the people in Iquitos live. They don’t live up in where the tourist hotels are. It really begins here. Lovely place, not particularly good at housekeeping. This is the plaz de armes in Iquitos. This is where the restaurants are and so on. Of course, most of you know that Iquitos is now the epicenter of the world ayahuasca tourism phenomenon for good or ill, and lately it’s possibly for ill. That’s a whole other question that Dr. Fogarty addressed. e were really involved in that. That wasn’t the reason we were here. We were here because we had the resources of Juan Ruiz and the university to rely on. But you can go to the marketplace in Iquitos and buy all these herbal remedies. This is what people rely on for their basic health care. This is the Amazonian ethopharmacopeia, if you want to call it that. A lot of these have not been investigated, really. They’re used, but there’s not that much work on their chemistry. Interesting, based on what Jean Mar said, there is certainly in this marketplace a preponderance of aphrodisjaks.

[00:35:19.140]
That was another thing that stood out. Afrodisjak is a very widely applied category to herbal remedies. If there’s any psycho activity or stimulant activity at all, you can pretty well bet that whatever else it might be, it’s considered an aphrodisjak. This is typical herbalist stall in the Kitos. Barks and tincters and crude extracts, and not all plants either. Extracts of snakes and turtles and other things you probably don’t want to think about too much. Dolphins’ penises, for example. Another valuable resource for this study was the U NAP Herbarium. That’s Universidad National Amazonia Parihuana. They have a wonderful herbarium. It could be better, but it’s a great resource. Actually, the Missouri Botanical Garden at some time put a lot of money into this herbarium, but that was 20 years ago. It needs to be bumped up. It needs to be improved. But it was useful for us because Juan Ruiz was the director, and we could work out of the herbarium. This is the man himself. I probably mentioned him. I’ve worked with him ever since I came to the Amazon for the first time in 1981 as a graduate student. We’ve stayed friends ever since.

[00:37:00.300]
He is an embodiment of this idea that when a medicine man dies, it’s as though a library has burned to the ground. He is not a medicine man. He would never call himself a medicine man. He’s a botanist, and he has one foot in science and one foot in traditional medicine. His father was a traditional healer, so I have not been able to show him a plant so far, even just a leaf or a fragment of a leaf. He can usually figure out what it is in a few minutes. I’ve rarely been able to stamp him. Sometimes he has to go look in a book. Usually, he has it on the tip of his tongue. This is a guy whose knowledge needs to be preserved because like the rest of us, he’s getting up there. I pleaded with him to write this stuff down, and he said, I don’t write. Why should I write? It’s all up here. He just doesn’t have that mindset. We’re going to work on that, see if we can get some resources, really, to document what he knows. This is our team, one, and then Alan Shumaker, who is an ex Pat living in the Kitos, and his wife at the time, Mariela Noriega.

[00:38:21.650]
They were very helpful with logistics and all that. Then we went to the field. These are two interesting plants. Well, it’s interesting the folk use that is noted on these plants. I don’t know, I forget which was which, but on one label, it was given to old men who have forgotten how to speak. The other one was given to old men who never stopped speaking but don’t make any sense. Back in the Bush administration, when this work was done, I was saying, Yeah, we got to fast track this one. But again, it’s just those fragments of information that are the key that may… Maybe there’s something to this. That needs to be looked at. Here’s another one I mentioned the other day, Tiliostocca, Lansio Meta, and the Acanthace, very common little ground cover. Nothing particularly stands out about it, but they say if you take a tea of this leaf, of the leaves, you’ll have a three day psychotic reaction where you lose your memory and you lose your sight. Then it comes back. During those three days, you have a conversation with the plant. But of course, you can’t remember what it told you. Here’s another interesting, lovely plant, Noryncia.

[00:40:10.260]
I believe it’s in the Mark Gravy AC. I’m not sure about this, but it is Noryncia. Actually, I have to thank Dr. Plotkin for, many years ago, identifying this family. And he said, Look at the Mark Gravy AC. It’s very promising. That’s where you’re going to find your novel psycho activities. You remember that? I don’t know if that’s true. You could have been. It is. Yeah, I think so. And very poorly investigated family at that. We took a trip. We took several trips into the jungle by boat and otherwise. This was one of the first ones. We went up to the Arboretum at Hinero Herera, a couple of days up river by one of these jungle taxi, three decker boats. This was on the way to disembark to the dock. I got out of the car and immediately fell into a ditch. I got my initiation, I guess, and get on these boats and travel with the people. Eventually, after a day and a night on the river, we end up at the Hineyra and this is our group. My friend, Mike Macky there on the left, who tagged along Carlos Pabón and our intrepid guide, Richard Fowler, also known as Acku and Juan Ruiz.

[00:41:52.220]
We didn’t need so many people, but you know how it is. Interesting bunch of characters went out there. Carlos Pabón is from Puerto Rico. He looks like he could have come out of one of the gangs in East LA. He would fit right in there. He s actually a brilliant molecular neurobiologist. Akul, what to say about H a Kue? Well, he has had a long history, shall we say? I heard recently that he passed on, but he had been a guide in the Kitos area for years. It was typically ex special ops, ex this, been to Vietnam, done it all. His novel was Give Me the Coordinates, I’ll Put You There. But we never really called on those skills, but he was fun to hang out with. This is the EAP, or Arboretum, the Instituto Investigaciónes Amazonia Parijuana. It’s partly government and partly university. It’s an Arboretum, but that doesn’t mean the trees are… Some trees are labeled, but it’s just a section of forest that’s been set aside. We went out and our merry band. A lot of these trees are big. You have to climb up to the canopy to collect the leaves and so on.

[00:43:23.960]
That’s why you have graduate students, right? They get to do all the fun or dangerous stuff. When I attempted this, I probably got about three feet off the ground and gave it up. Then Juan is just there and knows how to collect these plants and recognize them. This is just a beautiful passion flower that we found. And interesting insects, too. The stick insects were quite common. And the ants there, these are large ants called the isula ants. And you don’t want to get bitten by one of those because it’ll put you down for about two days. They’re very poisonous, very painful. At the field station at Janero Herero, they had a place where we could take our specimens every day and lay them out, make herbarium specimens. It’s a very important thing to do. Then we made our way back to Iquitos, eventually. This is a good way to travel in the Amazon. You certainly get to know your neighbors. There’s no first class Upper Burth here, as far as I can tell. We’re back in Iquitos, and that’s sunset over the Amazon. This is the group of people that worked on this, Dr. Hoey from chemistry, Dr.

[00:45:04.810]
Roth, Juan Ruiz, Alan Shumaker, and Mariela, and then some technicians in Dr. Hoey’s lab that helped process the extracts. So thank you. Thank you for taking part.