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This session is the second installment of the 3 part spring, 2,022 progress in research webinar series.

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Today's webinar will feature 4 research projects developing novel tools to remove learning compounds from the environment.

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If you missed the first webinar on her and poly floral alcohol substances.

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The recorded archive is now available on the flu and website.

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The final session of this series will be on Friday May thirteenth, and we'll focus on plant and fungal-based bioremediation.

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With that we are now ready to get started first. We will hear from Dr.

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You name Tang and Dr. You. shaun. Dr. Tang is an associate professor in the Department of Civil and Environmental Engineering, in the College of Engineering at Florida State University.

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Dr. Shan is an assistant professor in the Department of Civil Environmental and Construction Engineering at Texas Tech University.

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You can read full bios for all our presenters on the flu and website. Dr.

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Tang and Dr. Shun, I will now turn it over to you.

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Okay, Thank you, Cindy. So let me share my screen

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Okay. So since Cindy for the introduction, so hello, everyone.

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So my name is Youachelson from Texas Tech University today.

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I'll be presenting with Union Town from further State University. So our an Ihs topic is enhancing of Groundwater co-contaminated by chronic monitor organic compounds and the one for dioxide using

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novel microsecondic materials. So first we have a big teams.

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So Dr. Town from Florida State University leads to students working on power limitations, and Dr.

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Josh How and I, these 3 students from Texas Tech University.

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So we are working on material scenes in the design. So Mr.

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Joe from Josin Tech will join us in the monthly regular meetings to discuss whether Project progress.

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So we're join him in the year for and the year 5 of the project.

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So for the senior devices we have support the front Geos in Tech and the Epa New Durand during Graves and the genie comments for the laboratory tests.

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We have support the front Saturday and the University of Illinois and Chicago's.

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So back to our problems. So our problem is originally from the ground, was her contaminations.

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So. So one for docs is a very common organic solver. that use the first stabilizer of Cboc's: So the first the problem is the concentration is really low.

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It's very difficult to remove the completely so because these 2 components coefficients with each other.

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So the cboc's existence will incubate one for the outcomes power degradation.

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So the so the problem is, These 2 components need to complete opposite environmental conditions for about degradation.

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For 1 4,000. They need aerobic and the 4 Cbocs don't need an aerobic.

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So based on the problem analysis, we propose a solution that made up to component.

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The first component is, we try to cultivate some bacteria that can efficiently decrease the one for the oxygen at the very low concentrations.

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The second component of the solution is, we try to develop normal microsecond excellence, to try to make 2 kind of solvents that can physically efficiently remove one for the oxygen and the cdosis based

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on the solutions have different aims. The first aim is needed by Texas Tech University.

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We are doing computational simulations Synthesis material calculation to screen some highly efficient zorbent that can separate one for dog scene and the Cboc's.

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The second aim is needed by for the steel andesitate.

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They're doing cultural enrichment isolation. and the post calculation to screen some bacteria that can efficiently remove one for that and very low constitution.

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When the goes of these 2 end are cheap, we are going to interact with each other in the year 3, and then we are going to test our material, enhance the power immunization.

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This concept, and after that we are going to Conductor Colin study at Geosy Tech.

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So basically our research is fundamental research. But we also cover a technology development.

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That's why we collaborate with geos in tech so let's back on to our the progress of our project in the year one.

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So first, we conduct the material design, the synthesis, the first.

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We conduct 3 achievement in the modeling part. So for the first a target that would try to identify the most abandoned also probable conformer for the our microseconds, because these studies have been seldom

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study before, for example, it's Peter 6 a ring microsecond molecules, because molecules can rotate can interact with each other.

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So we first find that the c 7 is the most abandoned conformer for the you know, 6 are ring, so we'll use it as a representative molecule in the later simulations.

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So the second part of the Us. probably the the problem is trying to, trying to probe the transport of the barrier relating the absorption in different tests of microcycles.

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So basically, you can see this is the microseconds. and we probe a variety of contaminants from the entrance to the cavity, and we see the energy.

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Change. So I don't know how wide this figure is shows up here.

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So then, based on these 2 problems, and then we analysis the material based on a strength and adoption of Cdoc and the one for dioxide in computer 6 array based on microseconds.

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And and then we can see the binding energies.

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We can see slightly more negative binding energies for one for the oxygen compared to Cboc, which means they are, and a generate more favorable to combine find the one for the vaccine.

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But this difference is marginal. Oh, this is the progress of our modeling inside it. So my group is also at the same time doing some synthesis in addition to Pd. 6 ring.

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We are also investigating some other microsecond basis.

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Solutions, such as several dection and a resource in array.

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They have different shape, and they have different chemical environments.

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We also use different across linkers to indo the the finals open with different chemistry, such as the neutrals, oven and non exorbitant, and a foreign assault.

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So for the first year we are synthesizing the with a second detriment based the market solvent, and also pillar ring base.

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That's already based on the synthesis so we also started a P. of 6 A.

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Ring. but we find that the sed activity is not that significant as indicated by the simulations that's why we're moving on the data central adaption basis solvent and this is also being modeled by our work in

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our group by Josh from texas tech and then we've seen this as 3 kind of solvent. the first open is anonic that's later choose over it neutral and then we find some interesting.

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Select activities so compared to a commercials over an active carbon in the Amazon, they actually have pretty high removal rate.

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When we dealing with very low pv level around 50 to 100 people level one for the housing and the Cboc's, but they have no selectivity.

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But for our solvent like it's open to one in the street, and we find a known removal for one for Doc scene, but that they have selected the mobile of Cdc.

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Although the remote rate is the restro, we are going to improve it.

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In our later studies in the adoption tree. we also see these kind of selectivities.

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So this is the progress of our specific in one let's move on to the doctor town to introduce the progress of specific in 2.

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Alright, Thank you. Rashell. Hello! everyone. The second specific Am.

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Is mechanistic. Understand your highly efficient work for the oxen, metabolism, culture.

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We have maintained this mixed the country in the lab for 3 years.

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We know that it can degrade one for the option at Ppd.

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Levels. Major hypothesis is that this mixed culture contained species that can degrade one for the oxen at low concentrations, and that they can be isolated.

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And the methods that we use to pull in Richmond and isolation are similar to the literature.

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For example, we use a centrifugation combined with solution.

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The major difference is that we use low one for the outside concentration, as you can see from the figure above.

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It's the concentration is 1 min event per liter or less.

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And in the end we successfully, in which the 6 cultures, as shown in the 6 bottoms resp., they can be through the divided into 2.

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The 3 bugers on the left originated from an endpoint of ditch and the 3 bottles on the right, on whichated from an activities large.

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There's a big difference in the color of the culture room for naturally chase, but ground for the activities large.

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The next step is to transfer the mixed cultures into auger plates so that we can isolate them.

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And we say a lot of isolated colonies. Then we further transfer the isolated economy colonies separately into controversies, and we attempt 54 isolated cultures, and we also tested the ability to degrade one

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for the oxygen in the culture bottles, and we conform to that.

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85% of the isolated conscious code degraded one for the whole dioxide.

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The next step is to find what this isolated conscious are, and we sequenced.

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The 16 is our Dna, and then plots the results in the this final genetic tree which is on the right side. And this we started with 5 cultures.

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And they are in the red circus. Some of the coaches are closely related to.

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Conscious that's unknown to be able to degrade one for the option in the literature.

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Yeah, some are not and could be. We will now go back to the other way.

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And then next question that we want to answer is, how good are they degrading one for the oxen?

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So we did that kinetics experiment, and we focused on that.

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We started with the the coaching, the wind supper, and then we did a a batch.

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Butter experiments monitor the one for the oxygen concentration which is shown in the pink diamonds, and we also monitored the mask concentration, which is strong in green to end us at the

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same time we developed a model based on the 2 equations on the West side, and by feeding the model we experimental data, we determined the kinetics parameters, which is new red, and and after getting this parameters we

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compute them with the literature we first look at the parameter K is, which is, they have maximum rate concentration.

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And how can we go to the next slide? Thank you.

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Yeah, the the results, the isolated culture in our study is in green, and the literature results are shown in blue.

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For few coaches, and our range for mixed coaches, for the half maximum concentration.

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The the culture that we isolated has the a case that is close to the lowest reporting in the lead, which I mean, that's this culture can reach its maximum growth potential at a very low one for the oxygen

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concentration that is a characteristics we were looking for.

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We also compelled some arithmetic parameters, and upon that this culture had the lower balance, decay.

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Wait high bio mess yield, and no maximum specific utilization rate, which is not surprising because we use the logo for the oxen.

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Concentration. So, based on this characteristics, we think it seems to be a good beat once without some degradation.

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At no concentrations. So we started out, like to make conclusions for the specific ends that we have already worked out specifically.

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And why focused on material research we got regarding modern group.

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We have simulated pillar or 6 ring, and found that the weakening selective one for the outsome over klobinetic compounds and our experiments conform these modeling results because it the selectivity is

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weak. so we switched to another at this moment later, which is better cycle.

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That screen. On the other hand, this at this moment short one selectivity towards combinated volatile organic compounds, when both work with the outs and and the crew and global native volatile panic compounds of

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descent, and then in the second specific, and which folks to own mobile research, we have

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In which the 6 coaches mixed approaches, and identify the 5 queue conscious and the characterized, the one pure culture which seems to be a good fit for degrading, more for the absent at low

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concentrations. Looking at our progress. this is a 5 year project.

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We are currently in early year 2, and we have 4 specific ends who are specifically m one.

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The material research. we have already synthesized material that can selectively absorb Cdcs.

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What we want to do next is to books on synthesize and material that can selectively absorb one for the oxygen for specific M.

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2 which focuses on microbial research we have already in reached the 6 mixed cultures, isolated 54 queue coaches, and the characterized one efficient queue culture and our plans to keep kept

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the rising more few cultures that we have already isolated.

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And it was We have that Mike. Well, Macrovia, queue culture and the materials.

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Then in specific, and we we were started out into actions and instructions. Pacific M.

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4 where we started the interaction in the columns.

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We start out with the like to thanks the Nihs for funding.

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And thank you all for your attention.

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Thank you. we'll now take questions for Dr. tang and Shan.

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Yeah. As I mentioned before, you may submit your questions using the Q and A. pod down in the lower right corner.

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Did you see one question here? How would this technology be applied at a site with both one for daxing and chlorinated compounds?

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Maybe I can take the first best shot that's supposed to talk about access c.

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To remediation. we could have 2 stages, so that in the first, in the first stage we put in a material that selectively at the clue, and it compounds, and that material will be supplemented with bacteria that

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can degrade the global native compounds and then because they did not at the zoom month for that accent.

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So one couldouts, and will be treated in the second stage.

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Again, we will use as not a type of material, combined with some balance to deep with them.

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So that means the 2 groups of contaminants are degraded separately, because, material, the selectivity of the material that was possible.

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Do you see that working the same way in an institute application?

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I think potentially, it could be applied to in c 2 applications.

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So that's the we inject the media the the attachment.

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The absorb at this opens and bacteria into the source source that we are.

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Take care of them

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We need to file within the So so. But the techniques will be slightly different, because we cannot inject the big absorbance into the subsequent area.

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We need to use smaller at this open, which is also part of our.

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Thank you. Okay, I'll just remind books to put the questions into the Q.

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A I have another one for you here so have you run into any intermediates from the one for the exact same biodegradation.

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Well, I I think that's it very important aspect of the second specific

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We we measure one for the option for in and it's intermediate to coach, that we have already isolated, and for the first few culture that we characterized, we did not see intermediate production and that conclusion is based on

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2 observations. First, we measure the chemical oxygen demand and phone. Did that?

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Yep. The chemical oxygen demand during the degradation was confuted by both for the outsing, which means no intermediates.

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If they existed, the invention for in concentration. and we also, use, too.

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Yes, from a talk with you, mess spectrometric to see we can detect any intermediates which from there.

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So So with the first culture there's no thing media production product we will see the other conscious.

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Maybe some of them can produce further investigation required alright thank you

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Let's see. So there is a I Guess okay yeah One of the participants asked.

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Absorption would not results in compound degradation.

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So what is your plan on dealing with absorbing material after absorption?

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Yeah. So that can take take the these questions, so that that actually is

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What do we do? right? So we developed from materials that can selectively remove both to compound the components.

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And then Docker Town develop a bacteria that efficiently degraded one for that scene.

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And then at the same time, that clear degreeing Cboc has been already well studied, so they have very much for the culture, and can decrease efficiency.

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Then in the Us. 3 when we 2 teams come together and then we'll have one material that can select remove one for theoxing, and then that will have bacteria attached on that and then to further, the degraded one

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for the outside, because it's a bacteria it's a it's a conjugate enriched the column, and we have another column has cbo singing reach that can develop the for a

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Cboc division. So we will integrate the hour to team together to to see the material enhanced about the equation.

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That's what's our plan yeah we hope to make results.

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You know, 30 year presentation like this one Great. Thank you.

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Have a couple more questions for you here. Can you talk a little about?

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How computer simulation interacts or helps with your material design experiments. Yeah.

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So that's a good question. So I think the selectivity is very important.

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So based on the computational simulations, they will tell us that, what's the like? a binding efficiency finding preference for these 2 contempt, because they're very very similar, as we mentioned that the p 0 6 we already

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see some study, we find that there are difference in marginal and we don't see a good selectivity.

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So then all also. So our team will also study the other microcycle ring.

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So if the ring doesn't really have a difference so then we will move on to the the like.

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A cross linker to show that what internal chemistry, like hydrophobicity will tell us.

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So based on their their their their side, and also based on our experiment side, because we will do parallel together right? and then we will try to select the best solutions that can efficiently differentiate the user to

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chemicals. Thank you. Okay, I have another question for you.

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So what separation efficiency do you hope to achieve?

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And the one for a dioxide in cdc mixture.

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That's also a a good question so so far we don't know to what's the detail because everything is in Tv levels.

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So, or we want to know, for example, so if we selectively remove the Cboc.

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So what's the left cboc concentrations won't be inhibit the one for dx integration.

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I think that's something we need to investigate so that's the key part key, important part.

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So we can leave some Cbc. left. but we don't want inhibit the one for the housing integrations.

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Great. Thank you. I I also want to add on the qu here on that separation.

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So the first thing is to separate in the end we hooked the macro license.

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We'll be able to deal with there will be no contaminants the

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Thank you. Okay, I do not see any more questions in the chat,

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Or excuse me in the Q. A. pod. So if anyone has any more questions, please put them in now.

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Otherwise Dr. Shan, or tang do you have any other comments.

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You'd like to make before we move. to our next speakers just want to thank everyone for. attending our presentation is webinar, and also who the questions Thank you.

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So stop the sharing. Thank you. Okay, and if anyone has any further questions. Remember, you can put those in the Q and A.

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Pod, and we'll pass those along Okay, next we will hear from Dr.

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Lewis Sympony. Dr. Simprini studies biological processes for the treatment of hazardous waste, and on the fate and transport of organic contaminants in the environment at Oregon State University Dr

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Simprini, I will now turn it over to you.

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Thank you, and thanks for the opportunity for us to present our work today.

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Today i'm gonna discuss about development of passive sustainable co metabolic systems for the treatment of complex mixtures by encapsulating microbial cuts, cultures, and slow really substrates and

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hydrogels and my collaborators at Osu are Skip Roachford, who's doing material science and and cape and fog, and Mike Kyman, who keeps us honest on the

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microbiology that Oregon at North Carolina State University.

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So our our our first talk was a great introduction to my talk.

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We we out in the environment. We have very long plumes, for example, of one for Doc saying, that our costly to treat on and shown in the upper left.

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Here is the plume in Fort Carson that's several 1,000 feet long, that has one for docs and concentrations above 10 micrograms micro grams per liter, and so it's really very difficult to treat a whole

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plume those types of concentrations so one potential solution is creating passive systems for treating these plumes via the aerobic co metabolism and and And what we're what we're working on

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here is encapsulating pure cultures along with slow release substrates to drive, to drive the co metabolism in hydrogels, and then the contaminants with diffuse into the hydrogels to be degraded and the

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preliminary work on this concept was funded by by certain.

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So the microorganism that is doing the work for us is rotococcus rotacus, 2 1, 1, 9, 8 shown. here.

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It. It grows nice and orange on on a petri disk.

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Original studies were when you'd carry out co metabolism, where you stimulated this microorganism with growth on Isobutane as a gaseous substrate, it has a short chain alkene mono

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oxygen ace that initiates the oxidation, and here is shown the initial start of the Cov Metabolism of Cis, DCD.

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To an epoxide. In our work with the Isobutane we showed a broad range of chlorinated Alphatic hydrocarbons, and one-four could be simultaneously degraded by this

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microorganism my kimman at North Carolina state developed a fluorescent labeling method where we could label components of that short chain mono oxygenase enzyme and what's

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shown here. On the right is a labeling pattern, with growth of 2, 1 198 on different substrates and lane.

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One is shown growing on Iso Butane, very thick banding pattern here, but also when we drew it on to Buttewall, we also see abandoning a piano indicating that the model oxygenase was developed

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with growth onto Butanol, and and we also tested one.

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But well did not seal a banding pattern. But when we went over and looked at the transferation of one for Doc, saying, after growth on these 2 ox alcohols, we saw very rapid transformation of one for dx saying you

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have to growth on to Butanol, and then with some layout.

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Eventually we saw transformation, one for Docane on one, but no one, but all as well.

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So this brings up the concept of creating alcohols to drive the co metabolic transformation.

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The way we do that is, putting an orthosilicate inside our hydrogen, and an ortho silicate here is is has these oxygen bridges to molecules?

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That will then make the alcohol, and this would be a by just hydrolysis with water.

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It would slowly produce alcohols, over time so here we could encapsulate different orthos silicates to different to make different alcohols, and and we've worked with both tea boss here and 2 t boss that

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makes 2 butanol and 2 t boss hydrolyzes every rate about a factor of 20 slower than T.

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Boss. So on this technology you have some control of the rate of hydrolysis by the orthos silicate.

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You actually encapsulate? Well, getting back to the certific studies.

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What we did is we develop methods to ancapulate the microorganism and the slow release substrate in gelling gum beads, and this shows a column. pack.

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With these Joel and gum beads. Yeah. Early on in starting to flow groundwater that was contaminated with a mixture of Cis, Dc. E.

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1, one Pca. and one for Doc. say over time, You see this development on the right of the orange color throughout the whole column.

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This is 2, 1, 1, 9, 8 growing up in the hydrogen beads.

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Okay. So with prolonged stimulation, we get complete.

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2, 1, 1, 9, 8 throughout the whole column, with the gel and gum beads.

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And this shows the removal of the mixture of the 3 contaminants, and they were all at 250 micrograms per leader.

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And out here we're getting over 99% removal of all 3 contaminants with a hydraulic resonance time of about a half a day in the column.

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So this was kind of the proof proof of concept of this of this technology.

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So having that in our nihs prop a project we're kind of able to leapfrog and get into things we want to do in aim 3, and that's actually use the beads in a in a

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in a engineered system. and this is we created in the layout in a very large physical aqua for model, a funnel gate system and this shows the phone gate system.

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In here where we have the gates and the potential to create bio reactors, and then it's filled with sand, and over on the right it shows us packing them with yelling gum beads, So having the gelling gum beads in

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hand. Already we're able to kind of leapfrog into aim.

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3 experiments of creating a funnel, a phone and gate system to try and and treat the contaminants shown here in that system is the initial bio stimulation.

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So early. on these beads were we packed, liberated, one butanwall early on one.

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Butanol is liberated into the solution we're sampling. but then, with time one but Wall gets completely consumed.

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So the microorganisms are growing up in the gel income bees, and also show here.

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This is inside the reactor, the solid orange, so Don in red is the f one where we collect all the fluid coming out of the physical aqua for model, and we all we see but all concentrations decreasing

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with time, and on the right is dissolved oxygen.

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So the dissolved oxygen, decreasing with time, also indicates the bio stimulation of the beads within that reactor that we in place in the system.

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So we carried it out in in the in the in the system.

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Here we carried out some some tests with a surrogate, and a surrogate is a compound that will be co metabolized, and we could look at the apoxide That's formed so what we fed

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to the physical aqua for model is Isobutene, that is, co metabolized over to Isobutine oxide kind of a as a surrogate.

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Before we start going and doing experiments with chlorinated solvents in one for documents.

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And what we see here in the green is the buildup of the epoxide that is formed as we throw flow through that permeable reactor barrier that we created with the geling gum beads and then blue

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is the Iso, but teams shown to be at very low concentration. So we confirm co metabolism occurring in that bpack 150 days after we actually had in place the beads in that in that

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reactive barrier, and on the right is shown affluent concentrations from the reactor where we collect all, all, the all the f one coming out.

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And we see the epoxide in the F went, but at lower concentrations, indicating there might be fluid flowing around that funneling gate that may maybe not all the food is interacting with you that funnel and gate

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which is part of the design of our experiments to try and figure out if that is going on.

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We then moved on to a chlorinated solvent in the Pm.

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A 1, 2 Dichloro ethane. We did this because in the lab it has reaction rates around that of one for docsing

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But we wanted to test it out with a chlorinated solvent first. And what's shown here is in the brown on the bottom is the concentration in that reactive reactor in the funnel and gate and the blue

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is up grading of that. So the contaminant concentrations that would be mixing and flowing into that funneling gate.

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Right. And what we show is about a 60 to 70% removal.

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Okay of 1, 2, dye chlorine in this reactive system. the red shows collecting all the fluid at the end of the reactor.

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And what we're showing here is there is unreacted one to die.

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Clara Fay. There is fluid that is likely flowing around that funnel and gate funnel and gate system, and bypassing it.

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And this was also backed up by bromide tracer test.

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So what we have tests going on on the side of the funnel and gate, and in the funnel and gate, and the bromide tracer tests are in the funnel and gate and the bromide tracer tests are indicating that we

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do get flow around the phone gate, but the orange in the middle shows bromide going into the funnel and gate as well.

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So so a lot of work on, How do you actually conduct experiments in these physical aquifer models to try and figure out what is happening in these reactive systems?

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So moving on to the material science part of our project in the material size side, We're trying to develop stronger longer lasting a beads that we could mass produce easily and both in Co.

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Com encapsulate the slow release substrate and the the microorganism. and we're using design of experiments to look at the matrix of conditions you would study in developing these hydrogels So this is statistical

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method of trying to cut down the number of experiments you do, but also get a broad range of cover, so you could statistically predict which hat, which is the best jellen gumbee that you made over a range of conditions

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right. And here you put inputs into the model what we're dealing with is changing polymer concentrations and changing the time that we cross link to make the hydrogen beads, and then you have responses

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that you measure, and one is how strong out of the beads, how well they carry out co metabolism!

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What is the oxygen consumption that's occurring So that's a whole statistical basis for making judgments on how well you're making the hydrogen this just shows the process conditions our studies are in the first

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few are with deeds that are made of poly, vinyl, alcohol, and algernate.

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To make the hydrogels. We develop a system where we get all the components together in a series, and just with a simple syringe pump, we drop beads into a cross-linking solution to make the beads.

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And then we carry them out to do batch reactive experiments in the lab.

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And this just shows a simple procedure from making the 100 gel beats. right.

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So we're kind of the the momentums are up in here with a syringe. they're dropped into a beaker where they're cross-linked, and they make these beads that are shown on the

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right hand side over here. So in the design of the experiments what are we measuring after we make the beats?

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Well, one is we're measuring compression So we do compressing tests on the hydrogen beads to see how strong they are, and the blue is one day right after we make the beads and the

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Orange is 30 days after microbes have grown up in the beads in a batch incubation.

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They've been on a shaker table for 30 days shaking around. So we see that we do get we do get decrease in strength, which is we're trying to make these so they'd hold up for a long period of

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time. So one of the measurements over a broad range of experiments we conducted right And what we see down here is the billing to come metabolize this dce and actually that improves and gets faster with time from

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one day to 30 days, indicating microbes are actually growing up in the beads, and all the bees are capable of co metabolism.

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So we developed a process that the microorganism to survive. They're doing well in the 100 job beads.

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And then over on the right. here is showing oxygen uptake, and it's been an interesting observation, because we actually see more oxygen uptake at early time, and then at 30 days, and and that gives a question you know

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What is actually going on is this related to diffusional properties or things like that?

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And the beat. So a lot to try and understand when we fabricate and use these hydrogens and what you get out of the model when you need run these strange range of conditions is, you could actually make contour plots, of the results of

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the experiment. So this is the concentration of algebra. We put in the bead, and the concentration of B.

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Pva and the mixtures, and, as one would predict, the strongest beads are made when you have the highest polymer concentration, and as you go lower in polymer concentration, we the beads,

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are not as strong. What is also shown here in an opposite trend is oxygen consumption, and what we're saying is, we have highest rates of oxygen consumption on day one and beads that have the

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lowest polymer concentration and and we're trying.

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We're trying to think about what could be going on here, and it could be the matrix formed as you get more and more polymer in solution.

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You might be making a tighter matrix which is slowing down the rate of diffusion into the beats.

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So there's a lot going on here related to the material material science, and how we could actually test a beads.

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And again, the model is responsive into the measurements that you decide to make on the beats.

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Okay. So with that, we kind of going back to our columns again.

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So go with this we're going back. to our columns, and now these are the columns that are now packed with the new hydrogen beads that were were fabricating right and and and so these

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we have 2% by poly vinyl alcohol and allgenate, and we using tea boss that's going to liberate one.

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Butanol to grow grow the culture we have side ports to look at gradients in the system, and we in these systems as a supply of oxygen.

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We are adding hydrogen peroxide in the system.

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What is shown here earlier on, we saw a column that was completely orange with the gel and gum beads.

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Now we're starting here, you could see the orange color on the bottom of the column, starting to develop as we get into bio, stimulating, biostimulating the beads.

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So what happened? So we're looking at cisco in the column, and we're operated these now for about 90 per volumes of flow through the column, and what we're getting is over 95% removal

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of Cis Dce in the column, with a hydraulic resonance time of about a half day, with most of the activity coming in the very front of the call.

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We also see the production of the epoxide that we'd expect of Cisco here is early apoxide being measured, and then with time it's decreasing So what we're seeing is an increase in

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fish efficiency over time, and also the ability to combatolize the epoxide which is bringing is it towards complete mineralization?

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So we actually see out of the f one of the column the epoxide concentrations decreasing with time as well.

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One of the things we're seeing is that we don't have enough oxygen in the column, and we actually see one Butanol coming out the end of the column.

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So our rate of hydrolysis is fairly high and we've had to eat out hydrogen peroxide in order to consume consume the butanol.

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And we could see that here on the bottom you could see that we're running out of oxygen in the influence of the columns.

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So we haven't gotten stimulation, that nice orange color yet over the complete complete column that we have have here.

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So a little bit on side you know on beat optimization we've developed a fabrication method for a mobilization of both our our microbial culture in our slow release, substrate that should permit easier mass production of

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hydrogen beads in practice. this design of experiment method was successfully applied in the experiments to investigate different Polymer Co.

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Concentrations in different cross-linking times.

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And we pick the variables we're measuring in those tests a lot of tests to do.

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So one is beat compression, oxygen, uptake, coma tailism of the bees.

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Right. yeah on on as expected, the Pva Algernate hydrogen strength is correlated with the concentration of the polymer.

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So you go up in strength the higher the concentration of the polymer in the system on the beat application.

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We, You know, we have a proof of concept on the funnel and gate system using a big physical aquifer model to create a reactive barrier.

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We did this with our original gel and gum beads to see how well they would work in that funnel.

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Engaged system. We've seen Co metabolism of our surrogate Isobut team and 1 2 Dca.

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And this has been observed one year after the John Gong beads were put in place in the reactor.

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So coma tabloid has been going on for a year within the reactor.

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But we do see bypass issues that we need to understand a better through tracer tests and modeling of that system.

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And then we're now getting to test in in the columns.

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Our new made hydrogen beads right now we're looking at Coma tassel call metabolism of sisters to E.

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We will move up in the columns to one for Docane and other mixtures of contaminants in the flow through columns in other types of be B 4 relations and conditions.

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But right now we're getting over 95% removal of the Cis Dce, and we haven't even stimulated the whole column.

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Yet of the jelly gum jell and gumbeads. with my that i'd like to thank God.

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People working on the project skip, rove church and Caitlin fog, doing the all the hydrogen fabrication experiments and the doe modeling mohammed disease in helps out putting that

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big physical opera for model together and getting it running. Mike Hyman is working on the microbiology of North Carolina State along with Christie Smith, and then our trainees.

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Connor Harris is the PHD. student that has done all the hydrogen development and work with the doe model.

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A Jacob workcolor has operated the big physical aqua for model with the funnel and gate, and sean.

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Conway is working on the column study that I talked about at the end of the experiment, and with that I'd like to thank an Ihs for the funding funding opportunity.

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And if we have time for some questions, Thank you, Dr. Simpson.

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Yes, we do have time for some questions, and we have a few for you here.

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So first question, and i've heard this about prb is all over the place, So it's not just your experiment.

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How do you avoid flow around that barrier yeah that's a good question.

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That's why we actually set it up that there's room around the funnel and gate for the flow to go around it.

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So if we look at the construction we left the apps to the side so that flow, you know, if we did get it, It's a big thing to think about, you know, clogging of that permeable reactive beer over

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time, and and would you get flow short circuiting and going around the funnel and gate?

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And that's what we actually looks like was going on in our first experiment, and that's with trying to create hydrogen beads that are really robust right when you pack them in there.

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They won't disintegrate they would maintain their shape, and hopefully, you know, maintain their permeability.

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But but that's a really good question and that's Why, we're trying to do experiments in that physical model to to try and look at that excellent Thank you got a couple of questions about oxygen so how was

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the treatment affected by a decrease in dissolved oxygen.

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Yeah, what happened there is we're not seeing right now.

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Even when we analyze the ports right and have time to show that data on the on the column, the 3 ports we see most of the activities in the first half of the column, and then we run out of

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oxygen, and we're not getting stimulation of the beads in the upper upper half of the column right, and in that first experiment the rates of oxygen or the rates of

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hydrolysis has been high, higher than we saw in past column tests with the new beads, and so we've had the add hydrogen peroxide.

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In past experiments with prolonged use of the column.

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Rates of hydrolysis have dropped off and we're able to just use dissolved oxygen.

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But right now we're having to feed hydrogen peroxide as additional source, you know, in the system, and it is trying to develop systems that really consume low rates of oxygen right in the real world, because you know if

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you want to do things in a passive mode you're gonna need low, low oxygen.

387
00:49:19.000 --> 00:49:32.000
You. Yeah, thank you. Now, one participant asked, or they mentioned that addition of hydrogen peroxide can form radicals which would help in contaminate oxidation.

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But the same time might harm your bacterial colonies.

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So what are your thoughts on the trade-off between containers, removal and low bacterial abundance?

390
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Yeah, that's a good one. So the interesting thing is you know the the mud.

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These microorganisms, you know their grandp positive they're pretty tough critters, you know, and they seem to be able.

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We're adding pretty high hydrogen peroxide and they've been able to tolerate that, you know, in in these systems.

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And and it is surprising because we've gone up at some pretty high hydrogen peroxide concentrations, and they seem to be able to.

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Now we don't know you know the gradient you know if ones are being harmed early on, say in that column versus versus later right?

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But they seem to be able to tolerate it. The other thing here is what could go to the the orthos silicate that makes 2 t boss hydrolyzes at a rate of factor 20 lower than that t

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boss. so you would bring down the whole rate of microbial activity by this slower rate of hydrolysis, and you would likely, in our earlier column studies in the startup project.

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We didn't even have to had a hydrogen peroxide when we're looking at a slower release substrate in the system.

398
00:50:51.000 --> 00:51:02.000
So you also have control of this the slow release substrate that you choose, and the rate at which they hydrolyze, because that's based on the structure of that work, though or those silicates.

399
00:51:02.000 --> 00:51:10.000
Hmm. thank you, let's see I think we've got a related question here.

400
00:51:10.000 --> 00:51:17.000
You might have answered some of that already. But what do you think is the sweet spot for dissolved oxygen?

401
00:51:17.000 --> 00:51:21.000
And how would you amend that dissolved oxygen in the field?

402
00:51:21.000 --> 00:51:27.000
Application. Yeah, that's a good question. I mean you could think of this type.

403
00:51:27.000 --> 00:51:37.000
1 one is that we have to think about that there's diffusion into the beats right. and so whenever you think about the fusion you think about gradients of oxygen.

404
00:51:37.000 --> 00:51:41.000
You know we see this in biofilm studies that we do right.

405
00:51:41.000 --> 00:51:46.000
And so you really need that oxygen in the bead and So we're measuring the bulk solution of oxygen.

406
00:51:46.000 --> 00:51:52.000
But then it's a question what is the actual oxygen inside that bead I'm.

407
00:51:52.000 --> 00:52:06.000
Starting just by empirical evidence coming up with that you you'd like to have around 2 milligrams per liter of dissolved oxygen out in that bulk solution in order to account for diffusion of oxygen into the beads and and

408
00:52:06.000 --> 00:52:10.000
you almost get back to typical things. we look at in wastewater treatment, right?

409
00:52:10.000 --> 00:52:18.000
You want to maintain oxygen levels? Okay, at a certain level in order to get diffusion into flocks and things like that.

410
00:52:18.000 --> 00:52:32.000
So I. So oxygen is critical. We could also think, you know, and maybe as we go along here thinking of the slow release forms of oxygen, the solid release forms of oxygen in combination with the beads to try

411
00:52:32.000 --> 00:52:42.000
and make this a completely, you know, passive system. Okay? And so that is a potential study in the future.

412
00:52:42.000 --> 00:52:47.000
Great. Thank you. Okay, I think it's time for us to move to our next speaker. So thank you again.

413
00:52:47.000 --> 00:52:58.000
Dr. Simprini, and we will go on now to Let's see. next week we will hear from Dr.

414
00:52:58.000 --> 00:53:02.000
Tim Mattis and Dr. Wenching Sue.

415
00:53:02.000 --> 00:53:07.000
So Dr. Metis is a professor in the department of Civil and and excuse me.

416
00:53:07.000 --> 00:53:11.000
Civil and environmental engineering at the University of Iowa. Dr.

417
00:53:11.000 --> 00:53:16.000
Sue is associated professor in the department of Civil and Environmental Engineering at Vn.

418
00:53:16.000 --> 00:53:22.000
Numbers University. Dr. Madison, Dr. Sue, I will now turn it over to you.

419
00:53:22.000 --> 00:53:27.000
Okay, thank you. So let's see alright well thanks for the introduction.

420
00:53:27.000 --> 00:53:39.000
And here is the title of our of our project and it's been it's been a long title, but really what it's when it boils down to is that it's materials and microbes working

421
00:53:39.000 --> 00:53:44.000
together, which is in the spirit of this whole webinar series with chloride solvents.

422
00:53:44.000 --> 00:53:48.000
So i'm the pi to mattis and Winston sue is the Coppi.

423
00:53:48.000 --> 00:54:03.000
It's collaborative effort between the university of bio and Billanova and going to be acknowledging a funding from Nihs superfund research program here upfront

424
00:54:03.000 --> 00:54:11.000
And so the purpose of this project and the problem that we're addressing is, first off the clerk aquaria team.

425
00:54:11.000 --> 00:54:24.000
So pce and tce that's a you know These are common crown water contaminants, and in sits you aneroid by remediation of these compounds is fairly widespread utilizing organo

426
00:54:24.000 --> 00:54:28.000
halide, respiring bacteria, or O. Hrb.

427
00:54:28.000 --> 00:54:33.000
And so many of you probably seen this pathway before. This is the full declaration pathway of Pctc.

428
00:54:33.000 --> 00:54:40.000
All the way through, Cis Dc. Violet Chloride, and to Ethi, which is the desired end product.

429
00:54:40.000 --> 00:54:47.000
But as it's pretty well known that the limitations of this approach is the accumulation of Cis, Dc.

430
00:54:47.000 --> 00:54:52.000
And vital chloride in the field there's variety of reasons for that.

431
00:54:52.000 --> 00:55:04.000
You know bio augmentation is often done for, you know, in the case of there might not be the right organisms present, and there also could be other things like toxic co contaminants.

432
00:55:04.000 --> 00:55:13.000
Chloroforms just showing as an example here. Many of you who know me like we've been I've been working this problem for a while, but actually from the aerobic science that's kind of interesting to come in from

433
00:55:13.000 --> 00:55:22.000
the aerobic side for once and and part of the reason came interested was for some of these reports of what we call what we're calling pyrogenic carbonations.

434
00:55:22.000 --> 00:55:32.000
Matter. Pcm: Just have a picture of this. things like activated carbon coil activated carbon biochars showing some province and promoting Ohrv.

435
00:55:32.000 --> 00:55:44.000
Driven by irradiation. A lot of these reports are mostly anecdotal, but we're interested in how these materials are and their property specifically are shaping the microbi community interactions that can lead to

436
00:55:44.000 --> 00:55:53.000
enhanced by radiation. so that's what we're doing in this project is doing, you know, microbialology and material science.

437
00:55:53.000 --> 00:55:56.000
But the goal can developing Taylor, Pcm.

438
00:55:56.000 --> 00:56:10.000
That ultimately will improve halogenic organic pollute environment, outcomes, and some of the hypotheses that we're working under are that these materials are somehow improving my curvature interacting with my urban communities

439
00:56:10.000 --> 00:56:18.000
and positively influencing dehydration another idea that we're working on is taking advantage of the absorbed of capabilities of Pcm.

440
00:56:18.000 --> 00:56:24.000
That could be used to maybe sequester away, or calculate, or come, or otherwise inhibitory compounds.

441
00:56:24.000 --> 00:56:28.000
For O Hr. V. quick primer on what? Oh, Rb.

442
00:56:28.000 --> 00:56:38.000
Are doing they use in in the environment they use reductive the halogase enzymes called rdasis to catalyze clarity at the clarity thing declaration.

443
00:56:38.000 --> 00:56:50.000
I'm showing a picture of a cell schematic here, a lot of ohb use hydrogen and electron donor, and then they shut all those electrons over to the reductive halogenase which contains a a

444
00:56:50.000 --> 00:57:00.000
coronite cofactor that catalyzes that chlorination, and so in bacteria that are doing clarity, that the new coronation are 4 main ardaces.

445
00:57:00.000 --> 00:57:03.000
Pca. Tca. Vcr. and Bbca.

446
00:57:03.000 --> 00:57:07.000
And Pca. converts. Basically, Pc. Tc.

447
00:57:07.000 --> 00:57:18.000
Tca. can take tce all the way through Sis Final and to Ethane. and then Vcr and Bbca are kind of the main vonal chloride reducted the halogenases.

448
00:57:18.000 --> 00:57:30.000
They can convert Cis Dc and bottom for it to ethnic. And so that's the idea here is promoting a complete decoration to Ethane. and this is a study we're working with a culture a mixed

449
00:57:30.000 --> 00:57:34.000
culture called Stc. 9, The PCD chlorine culture it's commercially available.

450
00:57:34.000 --> 00:57:50.000
You can. sometimes comes through Paul hatsing or at them. it's where I get these this culture from and used environmentation frequently, and there was a study a couple of years ago with some meta genomics and proteomics showing

451
00:57:50.000 --> 00:57:56.000
that this culture is the main and hr b is the helicopters about 15 to 20% dealing with coities.

452
00:57:56.000 --> 00:58:02.000
There's also a desal photo bacterium present that does some of the work in this phylogenic tree.

453
00:58:02.000 --> 00:58:08.000
On the left is a showing different rdaces in the culture.

454
00:58:08.000 --> 00:58:16.000
So there are some Pca ardaces which looks like is possessed by the Desolfito bacterium, and then the health equity species.

455
00:58:16.000 --> 00:58:20.000
There have the enzymes vcra and Tc.

456
00:58:20.000 --> 00:58:28.000
Ea, so that's what we're doing and so Then the idea is that, you know, back with these materials that we are interested in.

457
00:58:28.000 --> 00:58:35.000
So there's some products out there it's a coil activated carbons, powdered activated carbons, maybe even biochars.

458
00:58:35.000 --> 00:58:43.000
We were looking at the interactions of Biochar, a popular bioterror which happened to have around the lab with this Stc.

459
00:58:43.000 --> 00:58:47.000
9 culture. So when we get it it's actually pretty happy and it it's making Athene.

460
00:58:47.000 --> 00:58:52.000
But it doesn't take very long it's actually grown often in a chemostat type type of situation.

461
00:58:52.000 --> 00:58:59.000
And then we start working with It in batch but on the left hand side here when there's no biochar present just in a liquid culture.

462
00:58:59.000 --> 00:59:07.000
It's not that difficult to get the culture a little bit unhappy, and just start accumulating Cisco, and starts making bottle chloride.

463
00:59:07.000 --> 00:59:21.000
Maybe after about 40 days or so but so what's interesting though we found was that when you pick the same culture, and you incubate it in the presence of this popular via char I don't know if it's anything special about

464
00:59:21.000 --> 00:59:28.000
the popular feedstock, or anything like that. but it just it's able to move past this Dc.

465
00:59:28.000 --> 00:59:38.000
And form completely declaring everything to athens so we thought that was an interesting observation, and then it's really not the material doing it.

466
00:59:38.000 --> 00:59:48.000
It's materials and the microbes works with together and that's kind of illustrated by some of these results where we're looking at some biomarkers in the culture for dehyde koi 16

467
00:59:48.000 --> 00:59:58.000
S. Vcra and Tca. genes and So in the cultures without Biochar, the the dialed Quadrants Aren't, growing so much up until about date. 40.

468
00:59:58.000 --> 01:00:05.000
Or so you saw a little bit of bottle chloride being produced, but in the cultures with bioterror there's several orders of magnitude more.

469
01:00:05.000 --> 01:00:09.000
The alquities with the genes Vcr. and Tca.

470
01:00:09.000 --> 01:00:16.000
That can do this complete conversion of Cdc to Athene, and in the we also extract the Dna from the Biochar.

471
01:00:16.000 --> 01:00:21.000
So they use. The helicopters are actually attaching to the biochar, and they have the Vcr and Tca gene.

472
01:00:21.000 --> 01:00:25.000
So again, this points to some interaction. and attachment.

473
01:00:25.000 --> 01:00:30.000
Direct attachment of the health equities to other, possibly other Lloyd.

474
01:00:30.000 --> 01:00:37.000
And that leads us to kind of the questions that we're working on in this project.

475
01:00:37.000 --> 01:00:46.000
And i'll have one ching talk to you about a one here in just a second, where about this tunable Pcm.

476
01:00:46.000 --> 01:00:57.000
Platform for synthesizing pcm like policymakers. So we're interested in zeroing in on specific properties of this of the materials that somehow lead to this enhanced fire mediation.

477
01:00:57.000 --> 01:01:13.000
And then, once we have these different materials, we can work with you know how they interact with the microbial culture ideally leading to, you know, finding out which ones promote more ethn formation versus others and then eventually moving on to

478
01:01:13.000 --> 01:01:25.000
aim 3 the development of a Taylor Pcm: that can be, you know, deployed in the field, and could also possibly be used for not just chlorinate Athens.

479
01:01:25.000 --> 01:01:32.000
But maybe other types of plurality compounds that are shown here in and sequest our way things that are not useful for the for the electricity.

480
01:01:32.000 --> 01:01:39.000
So with that I'm going to switch it over to Whatching, and she can take it from here.

481
01:01:39.000 --> 01:01:43.000
Thank you, Tim. Hi, Everyone i'm winching sue from Bolenova University.

482
01:01:43.000 --> 01:01:50.000
I'm the call pi of the project with the expertise in analytical chemistry and material synthesis.

483
01:01:50.000 --> 01:02:01.000
So as Tim just described in our preliminary results, Biochar somehow managed to accelerate the bio transformation of Pce all the way to Ethane.

484
01:02:01.000 --> 01:02:13.000
So the next question is, what is unique about biochar, and specifically, what properties of file char or pcm in general best supports the microbial network.

485
01:02:13.000 --> 01:02:24.000
To answer this question is very important, because it provides critical information for us to populate specific properties of Pcm.

486
01:02:24.000 --> 01:02:32.000
To enhance power remediation. So before I address that, I want to first talk about what is bioterror and what is Pcm.

487
01:02:32.000 --> 01:02:38.000
In general similar to activated carbon, which is a term that most of us are from with.

488
01:02:38.000 --> 01:02:43.000
Biochar also belongs to the family of Pergenic carbonaceous matter.

489
01:02:43.000 --> 01:02:52.000
Pcm. They are defined as the ink, solid, residual of the English combustion of fossil fuel and biomass.

490
01:02:52.000 --> 01:03:09.000
They have been traditionally used as passive absorbents, due to their large surface area high, poor volumes, and also the 8. polar surfaces, and I have included a little picture here showing the structure of activated

491
01:03:09.000 --> 01:03:14.000
carbon so contrary to the conventional wisdom.

492
01:03:14.000 --> 01:03:23.000
Recent studies suggest Pcm are actually reactive and They can promote bio transformation of certain contaminants.

493
01:03:23.000 --> 01:03:38.000
They're already different commercial products. for remedial efforts, for instance, clothes or activated carbon showing in this picture has been claimed to be able to enhance the bile remediation of

494
01:03:38.000 --> 01:03:44.000
hydrocarbons and chlorineet solvents in peered, reviewed literature.

495
01:03:44.000 --> 01:04:01.000
Both activate a carbon and biochar have been shown to transfer electrons from reducing agents in the environment to microorganisms or to facilitate the interspecies electron transfer among

496
01:04:01.000 --> 01:04:12.000
microorganisms. So the question is, what properties is importance, as you can show in this little schematic structure.

497
01:04:12.000 --> 01:04:20.000
Here it is postulated. This Kennon and hydrogen on functional groups in Pcm.

498
01:04:20.000 --> 01:04:25.000
Is responsible because it is a well-known Redox mediator, and it all.

499
01:04:25.000 --> 01:04:32.000
It is also hypothesized. The poly aromatic rain cluster, as shown in the second red box.

500
01:04:32.000 --> 01:04:40.000
Here can contribute to the electron transfer because they can potentially facilitate electron transfer at a faster rate.

501
01:04:40.000 --> 01:04:50.000
And both of those structures of Pcm. could also contribute to the electron exchange capacity of Pcm.

502
01:04:50.000 --> 01:05:04.000
However, the issue is all these properties. Surface functional groups, electronics change capacity, and the poly aromatic rain cluster which is reflected in conductivity of Pcm.

503
01:05:04.000 --> 01:05:08.000
Very significantly, depends on the type of the Pcm.

504
01:05:08.000 --> 01:05:18.000
So it really hinders us to understand which property contribute to the reactivity in enhancing bio remediation.

505
01:05:18.000 --> 01:05:23.000
Next, please

506
01:05:23.000 --> 01:05:32.000
Tip Next, please. well to identify which property of Pcm.

507
01:05:32.000 --> 01:05:40.000
Is importance. We have developed a Pcm. like Polymer, which i'll refer to as Plp to help us understand.

508
01:05:40.000 --> 01:05:49.000
The reaction mechanisms. it is important to notice that We're not really planning on pumping those Polymers in remediation, but rather we're using this P.

509
01:05:49.000 --> 01:06:00.000
At Plp as a platform to inform us what are the fundamental properties, we should be targeting down the line to tailor the carbon.

510
01:06:00.000 --> 01:06:04.000
So Let's talk about plp like pcm those Pcn.

511
01:06:04.000 --> 01:06:08.000
Plp Polymers also have large surface area and high microorosity.

512
01:06:08.000 --> 01:06:13.000
They're also highly conjugated and are aorphous.

513
01:06:13.000 --> 01:06:21.000
They also have superior affinity towards a series of polar organic contaminants.

514
01:06:21.000 --> 01:06:30.000
However, unlike Pcm. the attributes of plp can be individually tuned and made homogeneously throughout the polymer network.

515
01:06:30.000 --> 01:06:43.000
For instance, this shows the synthesis routes, and we can incorporate the quinone and hydrogen on pair which I described in the previous slide as a well-known radox mediator by selecting the

516
01:06:43.000 --> 01:06:47.000
R group in the linker through the synthesis process.

517
01:06:47.000 --> 01:06:54.000
We can also control the toology and the poor characteristics of Ppl.

518
01:06:54.000 --> 01:07:05.000
By selecting different monomers. For instance, the top one will result in a cross-length polymer versus the bottom one while resulting linear potomer.

519
01:07:05.000 --> 01:07:16.000
And so far, if you look at the figure on the right, we have demonstrated that plp has similar or comparable reactivity compared to different Pcn.

520
01:07:16.000 --> 01:07:24.000
Including graphite, which is a model Pcm. and Gac, which is a commercially available product that is widely used.

521
01:07:24.000 --> 01:07:31.000
The reactivity is quantified by the decay rates of model condemnance as shown below.

522
01:07:31.000 --> 01:07:42.000
Next, please. So in this project We're particularly interested in 3 aspects of the properties of Pcm.

523
01:07:42.000 --> 01:07:45.000
The first one is the conductivity, as I previously talked about.

524
01:07:45.000 --> 01:07:53.000
So we will first synthesis plp 0, which is highly conjugated, but it also has no functional groups.

525
01:07:53.000 --> 01:08:10.000
No, and it is not conductive. Then, using the plp 0 as a base polymer, we can tune the polytomatic ring size of the plp, which will result in pilot aromatic ring clusters with

526
01:08:10.000 --> 01:08:14.000
different size that should be reflected in different conductivities.

527
01:08:14.000 --> 01:08:26.000
So the second properties were interested is the quinone and hydrogenome pair, which can be incorporated through the selection of the R groups during the synthesis.

528
01:08:26.000 --> 01:08:38.000
The third aspects that we're interested is the surface charge which we will select the carboxylic acid group and the Connery Ammonium group to incorporate into the Piaki which will

529
01:08:38.000 --> 01:08:43.000
confer nactive and positive charge on environmental, relevant condition.

530
01:08:43.000 --> 01:08:55.000
So if you put all of that together, we have a very nice tunable platform that will allow us to study the impacts on bow remediation from each properties.

531
01:08:55.000 --> 01:09:01.000
Specifically Conductivity Quinn on hydrogen, on pairs and surface charge.

532
01:09:01.000 --> 01:09:09.000
Tent next, please. Alright, So so far we have synthesis.

533
01:09:09.000 --> 01:09:18.000
The 3 different polymers. Pp. 0, which is the highly conjugated and morphist polymer with no functionality, and it is not conductive.

534
01:09:18.000 --> 01:09:23.000
We then tuned the plp 0 into this Plp.

535
01:09:23.000 --> 01:09:29.000
700, which is shown in red. It has a moderate size of paradigmatic rain cluster.

536
01:09:29.000 --> 01:09:35.000
The third one we have as a plp-h which has quinone and hygieneal pairs incorporated.

537
01:09:35.000 --> 01:09:41.000
We then conducted different surface characterization techniques, and on the right.

538
01:09:41.000 --> 01:09:45.000
This is the results from our solid state and our result.

539
01:09:45.000 --> 01:09:57.000
As you can see, all 3 polymers has 3 peaks labeled in blue, showing up which is the evidence to support the successful synthesis of the conjugated network.

540
01:09:57.000 --> 01:10:08.000
Then, if you're looking at the first, the black line which is a plp of h, there are 2 additional peaks showed up at 1, 67, and 1 96.

541
01:10:08.000 --> 01:10:21.000
These are the evidences that queno and Hydrochlan pairs are successfully incorporated, and if you're looking at the red line here, which is a plp 700, it has a broad peak from 100

542
01:10:21.000 --> 01:10:35.000
and 20 all the way to 160, which is indication of the formation of polar dramatic rain cluster, and by decom eluding this peak, we can calculate the aromatic index which, is

543
01:10:35.000 --> 01:10:46.000
indicated in theta here, based on the number, is suggest that we have a moderate size of plp. a polynomatic ring cluster in this plp.

544
01:10:46.000 --> 01:10:51.000
It is also supported by the conductivity measured here.

545
01:10:51.000 --> 01:11:05.000
So now the next step is to look at look at the impact of 3 pl piece in promoting the microbial network growth, and i'll let Tim take over.

546
01:11:05.000 --> 01:11:15.000
Okay, Thanks. And so, having some issues slide, okay, so just to wrap up where you know, we've just gotten started.

547
01:11:15.000 --> 01:11:25.000
Now that we that there are these Plp polynomials available for checking out what's you know how they're gonna be interacting with the Scc.

548
01:11:25.000 --> 01:11:33.000
9 culture. We did start a couple in initial experiments with them, and in this case we had some popular biochar and some

549
01:11:33.000 --> 01:11:38.000
This Ppl. 0 and the plp 0 700 and we threw in some.

550
01:11:38.000 --> 01:11:42.000
Gac is kind of comparison and I think you know it's.

551
01:11:42.000 --> 01:11:54.000
We're just getting started. But again we wanted to make an important point. So in this case, with 40 milligram per year, pce the properties of these, some of these plp is that they can be highly absorbed in so

552
01:11:54.000 --> 01:11:59.000
in this in the case it's interestingly the popular bioterror is not very zorive.

553
01:11:59.000 --> 01:12:02.000
When you throw in about 40 milligrams per liter.

554
01:12:02.000 --> 01:12:05.000
Pc. you get almost all of it in the aqueous phase.

555
01:12:05.000 --> 01:12:09.000
It's not absorbing too much Jac as you can see you know, considered relatively absorbed compound.

556
01:12:09.000 --> 01:12:21.000
Let's take away About 75% of that pce whereas at about the same loading these plps at least the plp 0, and the Plp 0 700 are a lot more absorbed.

557
01:12:21.000 --> 01:12:26.000
In, so the less it is. And I think this was a question that was asked earlier in the webinar about some other materials.

558
01:12:26.000 --> 01:12:31.000
How do you know balancing the absorption capabilities of these polymers?

559
01:12:31.000 --> 01:12:34.000
And you know by extension any materials that are made.

560
01:12:34.000 --> 01:12:46.000
You have to be careful with that, because it absorption too quick it could lead to, you know, interfering with the growth of the orb, and it's also gonna complicate the analyses because if you get a lot of

561
01:12:46.000 --> 01:12:56.000
absorption It's a little bit hard to document the bio degradation, so that's where we're moving forward with looking at you know, making sure we get the loading right with some isotherms and and also you know

562
01:12:56.000 --> 01:13:04.000
there's the ability to luckily we can the tunable platform provides us the ability to also have some plp that are less sort of.

563
01:13:04.000 --> 01:13:08.000
And so the where we're going to continue so I think what's the way that we're going to end it.

564
01:13:08.000 --> 01:13:18.000
And I just wanna thank my research team here, the students that do all the work, and Jesse Wall was a postdoc and weigh loose out a PHD.

565
01:13:18.000 --> 01:13:24.000
Student in my group, and Shisha's Bond Pron was the Postdoc, and Han Cow is a PHD.

566
01:13:24.000 --> 01:13:29.000
Student in when Shings group and i'll Leave it there for some questions.

567
01:13:29.000 --> 01:13:34.000
Thank you for that presentation. we do have a few questions that have come in hopefully.

568
01:13:34.000 --> 01:13:38.000
We'll have time to get to all of them I think Dr.

569
01:13:38.000 --> 01:13:44.000
Sue actually was reading this first person's mind and answer this question on her first slide.

570
01:13:44.000 --> 01:13:51.000
But they asked if the Pcm. served as a substrate, or if it was simply an attachment site with a biological culture.

571
01:13:51.000 --> 01:13:55.000
That's a really good question. I mean We had that question when we were writing this proposal.

572
01:13:55.000 --> 01:14:00.000
That's exactly what we're hoping to understand using this plp platform.

573
01:14:00.000 --> 01:14:16.000
So if you think about this plp 0, which essentially provides the surface area for microbes to grow. But it doesn't have functional groups, and it doesn't have conductivity, So it will be able to let us see other answer that question

574
01:14:16.000 --> 01:14:24.000
specifically actually great. Thank you let's see so someone mentioned on Slide 9.

575
01:14:24.000 --> 01:14:27.000
They. They wanted to know why Gack was or the granular activated.

576
01:14:27.000 --> 01:14:35.000
Carbon was superior to the other 2 Pcms in the contaminant decay.

577
01:14:35.000 --> 01:14:41.000
Oh, is that the so the figure on the right so that's a really good question.

578
01:14:41.000 --> 01:14:47.000
We're quite excited to see that You Know Gak really performs quite well compared to others.

579
01:14:47.000 --> 01:14:52.000
It's not surprising if you compare gac to graphite, because graphite really is a model Pcm.

580
01:14:52.000 --> 01:15:06.000
It doesn't have much of a surface area it doesn't have a lot of functional groups, and it's not that necessarily surprising compared to plp 0 because that plp 0 has no functionality either so, If I were to

581
01:15:06.000 --> 01:15:20.000
guess it's like the functional groups, of gap is very different here in typical light is well known to have a mixture of different oxygenated phones, and they could be important in this case and Yeah, they're also you

582
01:15:20.000 --> 01:15:31.000
know conductivity that could play around that's exactly what we're trying to get into more details in this project delineate the contributions from each properties using this polymer.

583
01:15:31.000 --> 01:15:38.000
Network Great. Thank you. So here's a question about the popular Pcm.

584
01:15:38.000 --> 01:15:41.000
Have you tried using popular Pcm. that wasn't stored in your lab.

585
01:15:41.000 --> 01:15:45.000
Person is interested in know if it was because the Pcm.

586
01:15:45.000 --> 01:15:49.000
Had been stored, and maybe it had different microbes as results.

587
01:15:49.000 --> 01:15:58.000
So actually the pcm that the poplar was it's. you know it's actually paralyzed the looks stuff that we've been working with paralyzed in the lab but we all expect to have any specific

588
01:15:58.000 --> 01:16:06.000
microorganisms that would have survived that Pyrolysis procedure and the microbes that are involved are in this Stc.

589
01:16:06.000 --> 01:16:13.000
9 culture. So it's but we we can easily test out other Tcm.

590
01:16:13.000 --> 01:16:20.000
Materials that came from other parts of the lab, I guess or not, in our lab, other parts of the country or elsewhere.

591
01:16:20.000 --> 01:16:32.000
Yeah, And I want to just quickly jump on that because Tim and I are talking about, including some commercially available bioterror for testing down the line, because, you know, making sure whatever material.

592
01:16:32.000 --> 01:16:40.000
We're coming up with does have the commercial availability down the line is quite important, and we're aware that thank you.

593
01:16:40.000 --> 01:16:58.000
So when organic contaminants are sourceed by carbonaceous materials or seasoning carbon harmonious, I guess materials such as in micropers, the bio availability of absorbed contaminants,

594
01:16:58.000 --> 01:17:03.000
decreases a lot. So how do you improve the microbial access to those sort of contaminants?

595
01:17:03.000 --> 01:17:10.000
So. it's a good point and I think I was making that point there at the end of the presentation that some of these Pcm.

596
01:17:10.000 --> 01:17:23.000
Can be a lot more absorbed than others. so the this particular biochar that we're working with is not very zorphic, and so it provides it provides other. I guess benefits for the microbes while

597
01:17:23.000 --> 01:17:31.000
not absorbing, the more a way the contaminants that they need, so that they can be what's it can be buy by available to them.

598
01:17:31.000 --> 01:17:38.000
So I think that That's How we would improve it is look at how, what are the some of the properties of you know of the Pcm.

599
01:17:38.000 --> 01:17:51.000
Like polymers that we can come up with that are less sort of necessarily, but also provides a benefit to the microorganisms for time and transformation.

600
01:17:51.000 --> 01:17:56.000
Thank you let's see so do you expect the same results.

601
01:17:56.000 --> 01:18:04.000
With other halogenic compounds. it's possible especially from an an aerobics.

602
01:18:04.000 --> 01:18:09.000
Then a robot transformations of other housing any compounds I think that that's possible.

603
01:18:09.000 --> 01:18:13.000
But again it absorption the Zor, the absorbed characteristics of the Comp.

604
01:18:13.000 --> 01:18:19.000
How corruptive and chlorine, I think, are by comparison not as sort of maybe, as say Pcbs.

605
01:18:19.000 --> 01:18:24.000
Maybe alluding to the next presentation or other compounds like that.

606
01:18:24.000 --> 01:18:34.000
So I think you have to balance that by availability aspect with, what other surface properties can enhance the remedy or enhance to the alignation?

607
01:18:34.000 --> 01:18:45.000
Yeah, and i'll also jump on that a lot of it you know the the material side is really messy, because if you think about the carbon, it's very different from one badge to the other.

608
01:18:45.000 --> 01:18:51.000
Right. So one bow chart maybe perform very different kind of another commercial available source.

609
01:18:51.000 --> 01:19:01.000
They just it's hard to compare them and that's Why, you know, we're using this tunable system to really understand what are the keys here, and then down the line.

610
01:19:01.000 --> 01:19:11.000
We can potentially translate that into products to Make that something that's more consistent across the board.

611
01:19:11.000 --> 01:19:19.000
Thank you. Okay, we're almost out of time here? but One more question is, how would you apply the biochar and the Sdc.

612
01:19:19.000 --> 01:19:26.000
9 in a field application. Good question. I was expecting at least some some of these practical questions.

613
01:19:26.000 --> 01:19:38.000
I guess there's a couple of different ways and so there's the there's that colloidal if we're if we're thinking about a groundwater remediation scenario there is this colloidal putting the

614
01:19:38.000 --> 01:19:44.000
the the biochar in some kind of coil form that you could inject, along with the Stc.

615
01:19:44.000 --> 01:19:50.000
9, or then there's also the bio barrier approach if we needed to use a more granular form of it.

616
01:19:50.000 --> 01:19:56.000
It could be then put into a permanent or active barrier type of application.

617
01:19:56.000 --> 01:20:03.000
Those are 2 main ones. I can think of I don't know if lynching You have any other ideas on that one.

618
01:20:03.000 --> 01:20:08.000
No, this is good. Okay, I think we're out of time anyhow.

619
01:20:08.000 --> 01:20:12.000
So i'll i'll thank you for that and I did have some other questions for you.

620
01:20:12.000 --> 01:20:20.000
We'll pass those to you afterwards, but thank you again for your presentation, and next step next up.

621
01:20:20.000 --> 01:20:23.000
We'll be hearing from Dr. Upall Gosh Dr.

622
01:20:23.000 --> 01:20:32.000
Gosh is a professor in the department of chemical, biochemical, and environmental Engineering at University of Maryland, Baltimore County, just up the street here.

623
01:20:32.000 --> 01:20:42.000
Dr. Gosh, I will now turn it over to you

624
01:20:42.000 --> 01:20:51.000
Let me do. Can you hear me now? I can hear you I'm, seeing your presenter mode?

625
01:20:51.000 --> 01:21:01.000
Okay, So thank you. Everyone really excited to present the work that we are doing.

626
01:21:01.000 --> 01:21:15.000
And that's the title of our work looking at leveraging interaction with solid surfaces to announce the biomediation of halogenated compounds.

627
01:21:15.000 --> 01:21:29.000
So let me start with describing the team here so That's me and the question i'm at the chemical biochemical and environmental engineering department here at the University of Maryland, Baltimore County and Kevin

628
01:21:29.000 --> 01:21:40.000
is at the Institute of Marine and Environmental Technology, also part of the newest demand in Multimore County was sorry to interrupt you.

629
01:21:40.000 --> 01:21:48.000
We can still see your percent or most i'm sorry what we can still see your percent or most presented mode.

630
01:21:48.000 --> 01:22:03.000
So how? how what should I do here? Oh, I see if you click on display settings. setting.

631
01:22:03.000 --> 01:22:15.000
Up on the lower left. Oh, I see How did they do it?

632
01:22:15.000 --> 01:22:29.000
And then so on the upper left. if you click on display settings

633
01:22:29.000 --> 01:22:37.000
You can swap the view. Is that better? Yes, sorry about that.

634
01:22:37.000 --> 01:22:47.000
Challenges with 2 screens. So the the other participants are postdoc to associate, and that lead Dr.

635
01:22:47.000 --> 01:23:05.000
Natty Lombard and Prd student you think who just joined us last year fall, and we have partners on our water one enough today, and Matthew Snowbridge from our cadis, Who's who are going to help

636
01:23:05.000 --> 01:23:15.000
us think about taking the signs to the field that's that's a big component of the work that we are planning to do so.

637
01:23:15.000 --> 01:23:24.000
The good thing about going last in a series of presentations is that the problem has been quite well defined by the other speakers.

638
01:23:24.000 --> 01:23:34.000
So I will quickly mention that we are looking at one of the distinguishing aspects of our work is that we're looking at a large spectrum going from Chloro.

639
01:23:34.000 --> 01:23:42.000
It tends to polycarnate by phenols, and, as you know, they contaminate water resources, ground water, or surface water systems.

640
01:23:42.000 --> 01:24:00.000
My congratulations is usable there that there's tons of literature on the feasibility, but it's confounded by some of the slowest of the process, and issues with interaction with use office and I

641
01:24:00.000 --> 01:24:11.000
would say that the we are understanding today of the deployment process, and how it is impacted by surfaces is somewhat poor.

642
01:24:11.000 --> 01:24:20.000
We have a lot of empirical observations that need to be quantified accurately to be able to.

643
01:24:20.000 --> 01:24:39.000
We incorporated within large scale models for groundwater or surface segment systems, and in some cases I mean even proposed that engineering may have moved a little further than where the science is and the reason I see that is for

644
01:24:39.000 --> 01:24:51.000
a successful application of technology it's often very important to be able to describe the signs very quantitatively, so that we can optimize the engineering application.

645
01:24:51.000 --> 01:25:10.000
And that's where it seems to be a lot of gap and i'm i'm really happy that in I ehs saw this gap and supported several research projects to be able to address this issue and

646
01:25:10.000 --> 01:25:22.000
bring the signs up to where it needs to be in a very quantitative way to be able to inform these applications in the field.

647
01:25:22.000 --> 01:25:28.000
So the specific aims of our work involve quantitatively evaluating the effect on between properties and microbial deploy.

648
01:25:28.000 --> 01:25:33.000
And that's an important one for us the quantitative aspect of the description.

649
01:25:33.000 --> 01:25:47.000
We want to evaluate the materials unlocalized bacterial populations and develop innovative materials for enhanced by the mediation. And we're focusing on biocharks and looking

650
01:25:47.000 --> 01:26:02.000
at vouchers that are available, activated on mobile, and then also imparting specific properties, are altering specific properties of these biochars. to be able to look to an up or down specific characteristics.

651
01:26:02.000 --> 01:26:13.000
That we are interested in, and then in collaboration with that Kate is, we want to be able to predict field scale performance of these innovative bar amendment materials through advance site models.

652
01:26:13.000 --> 01:26:30.000
How do we incorporate the the understanding into models that people would normally use in the consulting field to be able to evaluate technologies as part of feasibility study for a superfund site for

653
01:26:30.000 --> 01:26:40.000
example. so to give you a sense of where some of the uniqueness lies in our work.

654
01:26:40.000 --> 01:26:46.000
We are and the approach that we are taking we have.

655
01:26:46.000 --> 01:26:54.000
We are focusing on this particular organism it's Df: one that Kevin's lab.

656
01:26:54.000 --> 01:27:03.000
And In subsequent work, our engagement with Kevin we have, explored the use of this organism for Pcb.

657
01:27:03.000 --> 01:27:10.000
Determination, and there is quite a bit of literature on the past work that has been done on this organism.

658
01:27:10.000 --> 01:27:19.000
It's isolated from the field. and can use a pure culture of a single organism to study

659
01:27:19.000 --> 01:27:35.000
The specific kinetics. quite accurately and that's one of the uniqueness The other important aspect that we are excited about by and and and and for that reason we chose this organism.

660
01:27:35.000 --> 01:27:50.000
Is that this organism can decorate a decorate a wide range of Pcb molecules going up to some of the Hexa chlorobi funnels all the way down to Tce and Pc and

661
01:27:50.000 --> 01:28:09.000
What that allows us is a large range of, or to observe the declaration process for a large range of fluorinated molecules from Tce, which might be in that kow range between 2 and 3 all the way up to log

662
01:28:09.000 --> 01:28:27.000
kw of 6 or 7, And that's an amazing range to to play with and observe some of these interactions with surfaces, and that also allows us to think about how these applications pan out in the subsurface system in

663
01:28:27.000 --> 01:28:30.000
the ground water system for chloro teams versus in sediment.

664
01:28:30.000 --> 01:28:36.000
Remediation. we've already used that cardinal materials activated carbons.

665
01:28:36.000 --> 01:28:43.000
As part of the delivery of organisms into sediments that are contaminated with Pcbs.

666
01:28:43.000 --> 01:28:51.000
And we have published on that in the past, and we have seen enhancement of the Declaration of Pcbs.

667
01:28:51.000 --> 01:29:01.000
In the presence of the carbon. The underlying principles and the kinetics involved is something that we have a lot more work to do.

668
01:29:01.000 --> 01:29:07.000
So some of the specifics it's a strictly it's a strict anrobe it's slow growing.

669
01:29:07.000 --> 01:29:15.000
And The The cell densities are typically small, and this organism requires either a D. self, Vibrio or Co.

670
01:29:15.000 --> 01:29:20.000
Culture, and Kevin will talk more about that in the latest slides.

671
01:29:20.000 --> 01:29:33.000
So the types of experiments We are doing involve setting up these microcosm studies, and and it's important to pay attention to what's going on.

672
01:29:33.000 --> 01:29:42.000
These microcosms. They look simple. batch reactors But there's a lot going on there, so we have could have the dissolved electron acceptor.

673
01:29:42.000 --> 01:29:56.000
It could be the Pc. at the Pcb. in that media which is not pure water, is, there are a few other things in that media, and then we might be adding a carbon, a c.

674
01:29:56.000 --> 01:29:59.000
Is solvent, a black carbon material to look at.

675
01:29:59.000 --> 01:30:10.000
The interaction with these surfaces now one of the questions is when you are looking at a larger ring of hydrophobicity, What's the right way to pro and measure changes over time?

676
01:30:10.000 --> 01:30:25.000
We don't know how to take samples of that media, and then filter the the organisms, because some of these compounds are difficult to measure directly through a filtration system, because they absorb to anything that it

677
01:30:25.000 --> 01:30:39.000
comes in contact with. So our approach is to take heads for some samples for the model components and then use a passive sampler to be able to measure accurately the fully dissolved concentrations of the electron

678
01:30:39.000 --> 01:30:52.000
acceptor over time, and we have done that in the past for Pcb: So the plot that you see in the bottom, and this is from password, looking at the kinetics of the declaration of pcb 61

679
01:30:52.000 --> 01:30:57.000
that's a tetrahedral by phenyl to try Chloro, which is Pcb.

680
01:30:57.000 --> 01:31:08.000
23 and that's the log concentrations in picomones over a time period of about 3 months there.

681
01:31:08.000 --> 01:31:24.000
And this is the first time anyone who was able to make these kinetics and express it as a function of well. It is all concentrations of Pcbs super low concentrations in the picom modes and

682
01:31:24.000 --> 01:31:38.000
subpoenas, and if you convert that to Nano Grams for leisure, the lowest one is less than a nanogram per leader. so that that work really address one of the big questions people had in

683
01:31:38.000 --> 01:31:51.000
the field. What is the reason why pcbs don't seem to go away in the field while we have in any segment that we go and look, we find decorated, so why don't they decorate and what we showed here, is that

684
01:31:51.000 --> 01:32:02.000
it's not really a concentration bottleneck but it balls down to is a growth bottleneck that these concentrations are so small that these organisms can decorate, but they're not getting enough energy

685
01:32:02.000 --> 01:32:14.000
to grow. So the steady state concentrations in the field are really small, and that was that we published on that, and that's by looking at the kinetics very accurately.

686
01:32:14.000 --> 01:32:22.000
We can make interpretations that allow us to understand what's going on, and then start thinking about intervention.

687
01:32:22.000 --> 01:32:37.000
So the intervention there for us was the engineering of being able to grow these organisms to hire concentrations and bio augment in the field to be able to address and get around this growth bottleneck so

688
01:32:37.000 --> 01:32:44.000
coming back to what we are planning to do in our work here is to look at Pcbs.

689
01:32:44.000 --> 01:32:48.000
And Pc. de declaration in these microcosms.

690
01:32:48.000 --> 01:32:56.000
You want to do that and characterize these rate constants very accurately in both estrine and freshwater media media.

691
01:32:56.000 --> 01:33:04.000
And We want to have precise enumeration of the cell densities of Df.

692
01:33:04.000 --> 01:33:14.000
One, both in the iqus phase. and the solid phase, and you'll see some of the work done to be able to do those measurements accurately.

693
01:33:14.000 --> 01:33:25.000
We have to do these measurements accurately. to be able to get solid kinetic information, because then we'll be looking at small changes in those kinetics.

694
01:33:25.000 --> 01:33:39.000
When we introduce materials. you want to be able to, accurately quantify the software capacity in these black carbons with them without the alterations that we are planning, and then that will allow us to model the

695
01:33:39.000 --> 01:33:47.000
true kinetics in the presence of solids, and then we want to link that to changes over time.

696
01:33:47.000 --> 01:33:51.000
So here is a schematic of some of the early work that we have done so.

697
01:33:51.000 --> 01:33:58.000
First thing we want to be able to do in these kinds of systems is to be able to demonstrate that if we don't add the organisms.

698
01:33:58.000 --> 01:34:03.000
Things are stable over time. So what you see on the left is for Pcb.

699
01:34:03.000 --> 01:34:09.000
61 that's that would go to Pcb. 23 in the presence of

700
01:34:09.000 --> 01:34:13.000
The Df. one organism, but if we don't add the Df.

701
01:34:13.000 --> 01:34:19.000
One, we are able to maintain those concentrations at levels going down below.

702
01:34:19.000 --> 01:34:27.000
Going down to the low piccom orders to thousands of people molars over 90 days.

703
01:34:27.000 --> 01:34:33.000
They remain about constant and the same thing on the right hand side for T. C. E.

704
01:34:33.000 --> 01:34:43.000
Going to dce that's what df one would do it's it's staying stable over the 50 days of measurements at 3 different concentrations.

705
01:34:43.000 --> 01:34:54.000
That are shown here. We are looking at a range of black carbon materials starting from biochars from different source.

706
01:34:54.000 --> 01:35:02.000
By biomass, and also 2 different kinds of activated carbons, one made from coconut shell and the other from call between us.

707
01:35:02.000 --> 01:35:14.000
Call. Those are the 2 largest volume activate carbon's commercially available in the market, and then we would be altering some of those properties to look at.

708
01:35:14.000 --> 01:35:23.000
Changing, for example, like electron accepting capacity or the surface area this option capacity.

709
01:35:23.000 --> 01:35:36.000
So we want to have a range of these materials that allow us to probe how the different properties of the materials, and then saw

710
01:35:36.000 --> 01:35:52.000
How the declaration activity of these organisms, and we are measuring these these these soften characteristics in in the media in which these experiments would be performed.

711
01:35:52.000 --> 01:36:00.000
The slide here shows some of the some of the early measurements we have done off the Declaration kinetics.

712
01:36:00.000 --> 01:36:07.000
So. one of the unique aspects of this organism is that, as I said, is that it can decline.

713
01:36:07.000 --> 01:36:12.000
Pcb. and also a Pc. We looked at those kinetics.

714
01:36:12.000 --> 01:36:17.000
So if you look at the plots here on the left for example, that's for on the left It's for Pcb.

715
01:36:17.000 --> 01:36:22.000
61 going to 23 we're looking at the accumulation rate of Pcm.

716
01:36:22.000 --> 01:36:29.000
23 in the media, and That's plotted against initial concentration of Ecb.

717
01:36:29.000 --> 01:36:32.000
61 that's a large range of concentration that we are looking at.

718
01:36:32.000 --> 01:36:49.000
And if you think about what's happening here is that as the concept initial concentration increases the accumulation rate increases, and if you look at the average first order, rate, constant it's about point, 0 2 or

719
01:36:49.000 --> 01:36:55.000
so that's per day on the right hand side so Notice the the concentration scale.

720
01:36:55.000 --> 01:36:57.000
It's nanomolos there on the right hand side that's pce.

721
01:36:57.000 --> 01:37:04.000
And those concentrations are millimeter. so 6 orders of magnitude, higher concentrations.

722
01:37:04.000 --> 01:37:14.000
But when we calculate the first order rate constant on 0 4 that's about a factor of 2 to 3 different from what we saw for the Pcbs.

723
01:37:14.000 --> 01:37:24.000
Both for the Australian media and the freshwater week so There's something universal about the rate call intrinsic rate constant, expressed in terms of the affiliate is all concentration.

724
01:37:24.000 --> 01:37:35.000
That's invariant. for pce and pcb so that's telling us something here, and we are trying to understand that a little better as we move along.

725
01:37:35.000 --> 01:37:44.000
So. those those rate constants are in whistle across the 6 orders of magnitude.

726
01:37:44.000 --> 01:37:53.000
So we are exploring that further. and then looking at now, once we have a good understanding of those rate constants without the carbon issue servants.

727
01:37:53.000 --> 01:38:10.000
Now we can proven within that same. We can look at all these different solvent materials all through those links. I'll let Kevin take over from here and tell you more about the biology of these organisms and some

728
01:38:10.000 --> 01:38:14.000
of that most aspects that are really intrinsic to our project.

729
01:38:14.000 --> 01:38:28.000
Kevin. Okay, So what we've been working on is normalizing the cells, so that, this is critical so that we're going to measure the effects of different carbons on the kinetics, we need to be sure

730
01:38:28.000 --> 01:38:37.000
that all the culture conditions are normalized. So so for example, when we transfer the cultures shown first panel it's imperative.

731
01:38:37.000 --> 01:38:41.000
When we do transfers we find that if We don't let it go to 100% declaration.

732
01:38:41.000 --> 01:38:46.000
You know we get better growth. the sales don't like shaking.

733
01:38:46.000 --> 01:38:58.000
But in order to make sure the culture is somewhat mixed and consistent throughout, we find we can do a small very low rate of mixing, and it doesn't affect the physiology.

734
01:38:58.000 --> 01:39:06.000
That much. And finally, these cells grow naturally in co-culture with it to self vibrio.

735
01:39:06.000 --> 01:39:11.000
So in order to eliminate any potential effect that's a self-aberial which doesn't declarinate.

736
01:39:11.000 --> 01:39:15.000
It just offers a growth factor to this, to the decorator.

737
01:39:15.000 --> 01:39:19.000
We found that we can use extract from the to self Vibrio, and then remove.

738
01:39:19.000 --> 01:39:26.000
We don't have to have an act there's a very real culture in co-culture with the with the decorator.

739
01:39:26.000 --> 01:39:44.000
So these are some of the growth aspects we're working on so we've got the growth conditions, so that they're going to be consistent between experiments and if you could go to the next slide no call do you have

740
01:39:44.000 --> 01:39:53.000
control. Okay, thank you. So one of the things we're doing is pce

741
01:39:53.000 --> 01:40:01.000
We found ways to use Pcb: by using a reverse of a passive sampler.

742
01:40:01.000 --> 01:40:05.000
Essentially, we can have a steady state concentration of Pcb.

743
01:40:05.000 --> 01:40:11.000
By using, absorbing it to a passive sampler, and then it deserts into the culture medium at a steady state.

744
01:40:11.000 --> 01:40:17.000
Concentration, depending on how much we equilibrate with the passage Sampler for Pce.

745
01:40:17.000 --> 01:40:21.000
What happens when you had pce it sinks to the bottom.

746
01:40:21.000 --> 01:40:31.000
And the concentration that actually goes into solution the dissolution rate can limit the rate of the organism which is looking at the true kinetics.

747
01:40:31.000 --> 01:40:38.000
So we're achieve steady state we've developed a method of just absorbing pce to a sand column.

748
01:40:38.000 --> 01:40:50.000
We can then circulate the medium to the sand column, and depending on how much pce or the rate of flow through the sandcom, we can maintain a steady state concentration of Pc in the initial

749
01:40:50.000 --> 01:40:53.000
culture. so that's one of the things we've been working on.

750
01:40:53.000 --> 01:41:07.000
We've got that setup currently so that we can grow the cells and equilibrium with a constant concentration the next slide

751
01:41:07.000 --> 01:41:24.000
The next thing is we need to normalize cell numbers in other words, we're looking at different concentration rates, different rates of activity in response to a particular carbon We need to normalize cell numbers in each experiment that to do

752
01:41:24.000 --> 01:41:33.000
that we've developed molecular techniques We Use 16 s quantitative Pcr with specific primers for 16 s.

753
01:41:33.000 --> 01:41:39.000
And we can quantify the number of cells based on this we've gone through a number of methods for extraction.

754
01:41:39.000 --> 01:41:50.000
Traditionally we're extracting from sediments We had to adapt this now to extract from cultures, and we've got now our we very reproducible high up a highly precise method for rapidly

755
01:41:50.000 --> 01:42:07.000
looking at cell numbers using quantitative vcr and you know we see variability between individual sampling is usually linked to the number of cells we find Pce we're getting about one times 10 to the 7 cells

756
01:42:07.000 --> 01:42:11.000
is maximum, and with Pcb. it's about an order of magnitude less so.

757
01:42:11.000 --> 01:42:20.000
Now we can addly measure the cells and normalize the cell numbers when we're doing the kinetic studies next slide.

758
01:42:20.000 --> 01:42:28.000
And finally, we've we scale up the organism, so that we have a a source of microorgan for these cultures.

759
01:42:28.000 --> 01:42:31.000
We maintain them at their highest density by semi.

760
01:42:31.000 --> 01:42:41.000
Right now, semi-continuous culturing in a 20 leader for manner. And then we can just tap off of that. Whatever cells we need, at whatever concentration we need.

761
01:42:41.000 --> 01:42:44.000
And we're gonna see adapt this with a sand column.

762
01:42:44.000 --> 01:42:49.000
So we have continuous constant concentration of Pc.

763
01:42:49.000 --> 01:42:51.000
For growing the cells. we can grow the cells on Pc.

764
01:42:51.000 --> 01:42:58.000
And also use them for the Pcb. experiments the sales adapt immediately to Pcb. and with Pc.

765
01:42:58.000 --> 01:43:02.000
We get higher cell numbers, so that's how we grow and maintain them.

766
01:43:02.000 --> 01:43:11.000
Next slide. Hi, Paul, let you take over sure. So I know we had.

767
01:43:11.000 --> 01:43:16.000
We are going past our allocated time. But here does a quick summer week.

768
01:43:16.000 --> 01:43:21.000
We have experimental conditions optimized for the studies.

769
01:43:21.000 --> 01:43:34.000
We have an interesting finding is similar intrinsic declaration. genetics observed for client compounds with hydrophobicity ranging 3 to 4 h of magnitude.

770
01:43:34.000 --> 01:43:42.000
We have started characterizing black carbon materials. and we are preparing them for the next phase of the work.

771
01:43:42.000 --> 01:43:46.000
So the next steps are timing these black carbon for specific properties.

772
01:43:46.000 --> 01:43:50.000
You know, option is one of the key properties.

773
01:43:50.000 --> 01:44:00.000
But then the electron accepting capacity and the charge, and so forth, and the functional groups will be looking at those as we go to the next phase of this work.

774
01:44:00.000 --> 01:44:07.000
Measure the tailored declaration kinetics on the killed by black carbons.

775
01:44:07.000 --> 01:44:12.000
Relate those kinetics to the physical chemical properties of the black carbons.

776
01:44:12.000 --> 01:44:28.000
And then One of the one of the aspects of our work also involves looking at native materials and looking at how declaration is in the segment environment, associate themselves with different types of particles.

777
01:44:28.000 --> 01:44:31.000
Do. they have specific affinities of certain types of particles?

778
01:44:31.000 --> 01:44:37.000
And what can we learn from that As we start developing these by many amendment techniques?

779
01:44:37.000 --> 01:44:43.000
And not just coming from a synthetic lab standpoint, but also learning from nature.

780
01:44:43.000 --> 01:44:47.000
And then finally translate the work using advanced site models.

781
01:44:47.000 --> 01:44:57.000
And I will end with this slide, showing pictures of all of us happy to take questions.

782
01:44:57.000 --> 01:45:02.000
Thank you for the presentation. I have just a couple of minutes here for some questions for you.

783
01:45:02.000 --> 01:45:07.000
So how would you completely biodegrade the Pcb.

784
01:45:07.000 --> 01:45:14.000
Use once that anaerobic stage is done, Kevin, would you like to take that?

785
01:45:14.000 --> 01:45:18.000
Sure what we do, and we're doing this in the field.

786
01:45:18.000 --> 01:45:22.000
Now we combine the the the halo aspiring.

787
01:45:22.000 --> 01:45:35.000
Mike part is in the arrow. with an arrow and they're both absorbed 2 activated carbon pellets, and because they invite, is not it's not as absolute because of benthic activity, you have

788
01:45:35.000 --> 01:45:39.000
constant changing of the redox potential. Both those are organisms are functional.

789
01:45:39.000 --> 01:45:44.000
And so what's happening is you have the Chlorinator breaking down the higher chlorinated Pcbs.

790
01:45:44.000 --> 01:45:58.000
And till you have conjunctors remaining that have unfly chlorines, and those conjures are then can be broken down by the arrow and completely mineralized through natural conservative organisms that

791
01:45:58.000 --> 01:46:12.000
appear that occur in the environment. Thank you. Okay. So we had a question about Slide 6, but they asked, How How did you measure the electron?

792
01:46:12.000 --> 01:46:18.000
Accepting. excuse me, electron accepting capacities, black carbons more specifically.

793
01:46:18.000 --> 01:46:30.000
Using what method? and under which conditions. So this was a

794
01:46:30.000 --> 01:46:33.000
Yeah, I have to go back and look at the specific method we use.

795
01:46:33.000 --> 01:46:41.000
I think it was our titration method that we used. Sounds good.

796
01:46:41.000 --> 01:46:48.000
Thank you. Now in these you seem to be looking at estuary and freshwater conditions.

797
01:46:48.000 --> 01:46:53.000
With particular sodium chloride, molar concentrations.

798
01:46:53.000 --> 01:46:57.000
Have you looked at any implications? with salt water?

799
01:46:57.000 --> 01:47:20.000
Intrusion in coastal environments. For example, Kevin, would you like to address the question with salt water for this organism, and how it can switch back and forth

800
01:47:20.000 --> 01:47:27.000
These studies were working, we adapt the organism, and then we consistently use the same.

801
01:47:27.000 --> 01:47:37.000
A similarity of the medium for all the experiments so for these we're focusing more on groundwater and freshwater concentrations.

802
01:47:37.000 --> 01:47:42.000
But these same experiments can be adapted to any salinity.

803
01:47:42.000 --> 01:47:46.000
And I I think we propose to do mostly fresh water.

804
01:47:46.000 --> 01:47:56.000
But You're question concerning saltwater intrusion the same experiments could be done with that in mind to see if salinity is also a factor.

805
01:47:56.000 --> 01:48:08.000
I I don't recall. if we've written that into the proposal. But that's you know, based on the methods we're developing here that could be done as well because your same organism can be used excellent thank

806
01:48:08.000 --> 01:48:15.000
you. Okay, I have time for one more here. So with microbial deplorinators.

807
01:48:15.000 --> 01:48:21.000
So why we found in the natural environment. Why are the natural declaration rates especially?

808
01:48:21.000 --> 01:48:26.000
For Pcb. is so slow. You want to take that up, Paul.

809
01:48:26.000 --> 01:48:38.000
Sure I can take that so. So that that was kind of the central aspect of the work we did a few years ago to look at.

810
01:48:38.000 --> 01:48:43.000
You know this conference room that you know wherever you look in the sediments with pcbs.

811
01:48:43.000 --> 01:48:48.000
You'll find these declarators but then you know the the Pcb.

812
01:48:48.000 --> 01:48:55.000
Releases into the segments that happen heat that you know maybe 60 70 years ago they haven't gone away.

813
01:48:55.000 --> 01:49:10.000
We still have them. And so when we look, when we look at the kinetics, we found that the user organisms, So So there has been a lot of different perspectives on this in the literature, and one of the dominant

814
01:49:10.000 --> 01:49:14.000
perspectives was that there is a there is a concentration bottleneck below which Pcbs.

815
01:49:14.000 --> 01:49:23.000
Don't decorate, and we are able to refute that with those experiments and results that I showed that Pcb.

816
01:49:23.000 --> 01:49:27.000
Declaration is measurable in this at the same rate constant.

817
01:49:27.000 --> 01:49:42.000
So if you go back to this slide here, so the slopes of those lines are basically telling you in that log scale the first order rate constants and a cost those 4 h of magnitude going down to less than a pico

818
01:49:42.000 --> 01:49:46.000
molar concentration of in the water phase of Pcb.

819
01:49:46.000 --> 01:49:54.000
61. The rate constant, is still the same. and so there is no concentration bottle like there.

820
01:49:54.000 --> 01:50:09.000
What's happening is that these organisms in the field using Pcbs as the electron acceptor at those low concentrations of pico molars are not making great living They are in a very very poor

821
01:50:09.000 --> 01:50:17.000
thermodynamic mish so, although they have the enzymeatic capability to decline it's not helping them in in their growth.

822
01:50:17.000 --> 01:50:24.000
So if you think about microbial ecology in those systems where you have an organism that hardly goes their steady state.

823
01:50:24.000 --> 01:50:31.000
Concentration is going to be really small so when Kevin goes and measures these in the segments.

824
01:50:31.000 --> 01:50:34.000
You get something like correct me if i'm wrong kevin like Tennessee.

825
01:50:34.000 --> 01:50:50.000
2 cells per Ml. of sediments, yes, and and in our last people what we showed is that at that level of organism, density in sediments with the kinetic rate constants that we have measured in

826
01:50:50.000 --> 01:50:54.000
the lab it would take sanctuary is for Pcs.

827
01:50:54.000 --> 01:51:07.000
To go away, and that's, indeed, what we are seeing in the Now, the thing that we can change, and which is that innovative part of what Kevin has been able to do, and i've been able to help with some of the engineering of it

828
01:51:07.000 --> 01:51:15.000
to the field is that I can break that growth bottleneck by bringing these organisms back to the lab, isolating them.

829
01:51:15.000 --> 01:51:24.000
And that allows me to grow them at maybe multiple orders of magnitude faster on Pc.

830
01:51:24.000 --> 01:51:32.000
In a reactor, get larger numbers of those organisms, and then I have to buy augment back in the field.

831
01:51:32.000 --> 01:51:35.000
Bring up those numbers in the sediments from tennis to 2 cells by Ml.

832
01:51:35.000 --> 01:51:39.000
It was something like tennis to 5 to 6 cells, but Ml.

833
01:51:39.000 --> 01:51:45.000
And then we start seeing changes in the Pcb. concentrations in the sediments in the time scales.

834
01:51:45.000 --> 01:51:51.000
We like to see those changes happen which is in the months, 2 years, and not decades to centuries.

835
01:51:51.000 --> 01:51:55.000
So I don't know if it answered your question completely.

836
01:51:55.000 --> 01:52:08.000
But that's basically what we are seeing it's really the goal growth bottleneck which is linked to the slow rate of you creation that we see for Pcb is in the segment environment.

837
01:52:08.000 --> 01:52:13.000
And we can break that by by augmentation. Thank you.

838
01:52:13.000 --> 01:52:18.000
Thanks for that answer. Well, this is all we have time for today, so I want to thank Dr.

839
01:52:18.000 --> 01:52:22.000
Goshen, Dr. Sowers, and all of our other presenters.

840
01:52:22.000 --> 01:52:29.000
But please stick around for just a moment. participants I have a few reminders for you.

841
01:52:29.000 --> 01:52:32.000
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842
01:52:32.000 --> 01:52:43.000
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01:52:52.000 --> 01:52:59.000
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845
01:52:59.000 --> 01:53:13.000
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850
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853
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