Reverse Respiration

[Ken Burke]

@dasaltemelosguy @Guppysnail    Any chance we could see this as a peer reviewed article some day?

[Guppysnail]

On 6/16/2022 at 1:30 PM, Ken Burke said:

@dasaltemelosguy @Guppysnail    Any chance we could see this as a peer reviewed article some day?

That is TOTALLY @dasaltemelosguy’s department.  

[Ken Burke]

I can see applicability and utility for cleaning aquatic plants of invasive snails.  Wonder if it could works on muscles as well as snails

[Guppysnail]

On 6/16/2022 at 1:38 PM, Ken Burke said:

I can see applicability and utility for cleaning aquatic plants of invasive snails.  Wonder if it could works on muscles as well as snails

As I was reading up on cleaning methods I seen one talking of submerging in VINEGAR ph of 2.5 for extended time the zebra mussels from a research organization that said it was affective for boating and fishing gear that could be completely submerged. I have no way of testing Reverse Respiration(co2 ph 3) on zebra mussels though. I instantly thought of @Streetwise and the native collected plants he never felt comfortable with even after a long QT.

Edited June 16, 2022 by Guppysnail

[Ken Burke]

@Guppysnail  exactly.  But also for other applications like agriculture or plants used for waste water treatment. 

[Guppysnail]

On 6/16/2022 at 1:54 PM, Ken Burke said:

@Guppysnail  exactly.  But also for other applications like agriculture or plants used for waste water treatment. 

That is a very good point. 

[Odd Duck]

On 6/16/2022 at 10:59 AM, OnlyGenusCaps said:

I see above I have been given credit that I simply do not deserve.  I have been a keen observer of this process, and in awe of the dedication shown by these folks.  That is all.  My part has only been to be excited by the advances and research happening. 

I feel much the same.  I took a look at a few pictures, made a small remark about the BBA looking like it was denatured, and next thing I know I’m getting credit for contributing far more than I truly did.  It’s something I’ve been only on the outskirts of and waiting with baited breath to see the results.  There were a few convos about assorted things and I was happy to proofread.  Are you kidding?  Of course I’ll review it for you!  (It got me in on the secret early!  😆).

By far the bulk of the work fell on @dasaltemelosguy and @Guppysnail since they carried the load.  They need to get full credit for all this hard work, reading, and boots on the ground labor plus keen observation.  @OnlyGenusCapsI’m just glad you noticed @dasaltemelosguy work posted on another, not so kind forum, and directed him here where we can all appreciate his hard work and his flat out genius!  I’m proud to have been just a very tiny part of this project.

[OnlyGenusCaps]

On 6/16/2022 at 10:44 AM, dasaltemelosguy said:

I believe that @OnlyGenusCaps cleaned a bunch of java moss for fry with a SodaStream and he said snails just fell out of it that he never saw.

Yessir!  I did run this on some X-Mas moss.  I just used some store bought seltzer (not club soda as that has salt in it).  I also sealed it in to let the pressure increase.  Worked great!  Wiped out the snails & I've never gotten algae on the moss since.  I also did this on my crypt spiralis and didn't introduce anything unwanted.  However, that tank is currently covered in hair algae, but that's a cultural issue, not a failing of this system.  Interestingly, the crypts didn't melt for me.  I don't think this is a particularly melt prone species, but still, that was a nice bonus. 

 

[OnlyGenusCaps]

On 6/16/2022 at 12:30 PM, Ken Burke said:

Any chance we could see this as a peer reviewed article some day?

I think at the very least, it should got to Amazonas Magazine.  As a minimum.  This is such great stuff, it need to get out there as best as possible.  I do think there are peer review routes for it as well - that's probably based on whether he wants to take on that sort of work load.

On 6/16/2022 at 1:26 PM, Odd Duck said:

I’m proud to have been just a very tiny part of this project.

I fully understand and feel the same!  @dasaltemelosguy's generosity and openness 100% embody the kind and community oriented ethos I believe @Cory said he wanted to foster here in the intro video.  Good guy.  Good research.  Tough to beat that combo!  (I hope he's blushing.) 

[gjcarew]

Very cool work. CO2-bombing is a well-known strategy for pest control in terrestrial horticulture, especially in indoor grow operations for *ahem* certain plants. I personally use garbage bags filled with CO2 to manage pests on my houseplants. I don't know why I never thought of using this method for aquatic plants as well!

This could go in a journal. Each separate experiment (on chymatous zones, pest control, algae, subsequent growth rates) probably deserves an independent paper with the whole abstract-method-results-discussion shebang, just to make sure it's reproducible.

You lost me here:

On 6/15/2022 at 12:25 PM, dasaltemelosguy said:

“Nothing and Something Create Each Other”

(from Chapter Two, Tao te Ching, circa 500BC)

The findings above were quite significant for us, as they helped to confirm our theories of how Reverse Respiration utilizes available energy, be it photonic or electronic, to act as an algaecide.

Although it takes countless forms, mass, light, heat, x-rays, all are ultimately just energy. Be it a rock or a bolt of lightning, both are the same stuff, a form of energy. What a mass or a form of energy is defined as is relative and depends on the observer. 

It is the scale that creates the reality. If we were at the level of an atom inside of the rock, we’d see countless energetic particles moving near the speed of light. If we pull back, it’s just a motionless, dead rock. What makes it a mass is not inherent but rather where we observe it from. But in truth, it’s just the same stuff as the rest of the universe: energy.

The universal equivalence of energy is why Reverse Respiration kills algae more effectively in the dark. When light is present, the energy to facilitate photosynthesis is provided from an external source of light energy, photons. However, in darkness, with no photons for an energy source, the nearest available energy is therefore not photons but rather, the electrons in the algae itself. The proteins and enzymes become highly unstable at a pH this low and easily lose electrons (energy) in this state which in turn destroys all of the algae.

Tell me if I'm understanding correctly: You're claiming that in the absence of light, chlorophylls are "attempting" photosynthesis but somehow oxidizing the surrounding material? What is the evidence? I'm not questioning the phenomenon of algae dying, but that seems like an unlikely explanation. It seems more plausible that given high co2 + light, algae can consume enough CO2 through photosynthesis to create local microclimates of higher pH than in the surrounding solution. This might delay or even stop the "pH shift" that denatures the proteins in the algae.

Edited June 16, 2022 by gjcarew

[dasaltemelosguy]

On 6/16/2022 at 10:38 AM, Ken Burke said:

I can see applicability and utility for cleaning aquatic plants of invasive snails.  Wonder if it could works on muscles as well as snails

Hi, that's a really good point. As more people use the technique we'll start to see it's limitations! But it'll ever improve the data as we see reports of successes and failures. 

To your point, I did find this. I never tested this, I've never even seen a zebra mussel! So this is 2nd hand info but these people kill zebra mussels by infusing the water with Marimo moss balls with CO2.

They say that soaking the Marimo's in water with 1/3rd to 1X the CO2 in seltzer, they saw 50% kill off of the zebra mussel in 24 hours and 100% in 96 hours.  RR could be extended without harm to the plants if the cycles were normalized after the initial treatment. Basically, if it's changed to CO2 by day and plain water at night. You could perform multiple treatments to extend the bath time as it's a normal plant cycle vs reversing the cycle.  

One really cool aspect of the whole thing was frankly a surprise was when the plant was moved out of the seltzer to plain water, the pH shock literally shattered the snail eggs and killed the algae. We were planning on incubating them to watch for hatching after seltzer, but they literally exploded upon removal from seltzer and being placed in water. 

This may apply to the mussels, but I really don't know. the study is open-sourced so it's free. This is the link: Zebras and CO2

Here's an excerpt:

Abstract Control technology for dreissenid mussels (Dreissena polymorpha and D. bugensis) currently relies heavily on chemical molluscicides that can be both costly and ecologically harmful. There is a need for more environmentally neutral tools to manage dreissenid mussels, particularly in cooler water. Carbon dioxide (CO2) has been shown to be lethal to several species of invasive bivalves, including zebra mussels and Asian clams (Corbicula fluminea). We evaluated the effectiveness of unpressurized infusion of CO2 for 24 to 96 h (100 000–300 000 µatm PCO2) at a water temperature of 12 °C on mortality, byssal thread formation, and attachment of zebra mussels. The safety of elevated CO2 to a nontarget native freshwater mussel (Fatmucket, Lampsilis siliquoidea) was also determined. Elevated PCO2 exposure induced narcotization and reduced attachment of zebra mussels within 24 h. Mortality increased with exposure duration and PCO2. An estimated LT50 (lethal time to produce 50% mortality) for fixed PCO2 ranged from 24 h at 275 000 µatm to ~ 96 h at 100 000 µatm. (Seltzer would be about 330,000) Exposure of zebra mussels to CO2 for 96 h caused 80–100% mortality at all treatment levels. Fatmucket juveniles survived all PCO2 treatments but burial and byssal thread production were adversely affected during exposure. Our results demonstrate that CO2 is a viable option for management of zebra mussels in cool water and may have less adverse effect for native lampsiline mussels than current-use molluscicides.

[dasaltemelosguy]

On 6/16/2022 at 3:31 PM, gjcarew said:

Tell me if I'm understanding correctly: You're claiming that in the absence of light, chlorophylls are "attempting" photosynthesis but somehow oxidizing the surrounding material? What is the evidence? I'm not questioning the phenomenon of algae dying, but that seems like an unlikely explanation. It seems more plausible that given high co2 + light, algae can consume enough CO2 through photosynthesis to create local microclimates of higher pH than in the surrounding solution. This might delay or even stop the "pH shift" that denatures the proteins in the algae.

My writing made that confusing, I'm sorry. I actually should have added the flame tests we made form evaporating the solutions that the plants were cleaned in. In brief, we saw simple, chemical reactions from the intense electron field of the carbonic acid. We saw (I may have the % wrong, it's somewhere in my notes) something akin to 40% more denaturing in the dark. So, what you said my very well occur at least partially in the light. What we found in the evaporates was largely carbonate salts from the algae Magnesium, some Sodium and some Potassium. I left a lot out as the piece became so long! But it's all available in the downloads in PDFs at the end. 

On 6/16/2022 at 10:38 AM, Ken Burke said:

I can see applicability and utility for cleaning aquatic plants of invasive snails.  Wonder if it could works on muscles as well as snails

I don't know though. That would be extremely cool though! It was performed as a full, factorial so it's more or less ready for such. 

[Torrey]

On 6/16/2022 at 4:31 PM, gjcarew said:

Very cool work. CO2-bombing is a well-known strategy for pest control in terrestrial horticulture, especially in indoor grow operations for *ahem* certain plants. I personally use garbage bags saturated filled with CO2 to manage pests on my houseplants. I don't know why I never thought of using this method for aquatic plants as well!

This could go in a journal. Each separate experiment (on chymatous zones, pest control, algae, subsequent growth rates) probably deserves an independent paper with the whole abstract-method-results-discussion shebang, just to make sure it's reproducible.

You lost me here:

Tell me if I'm understanding correctly: You're claiming that in the absence of light, chlorophylls are "attempting" photosynthesis but somehow oxidizing the surrounding material? What is the evidence? I'm not questioning the phenomenon of algae dying, but that seems like an unlikely explanation. It seems more plausible that given high co2 + light, algae can consume enough CO2 through photosynthesis to create local microclimates of higher pH than in the surrounding solution. This might delay or even stop the "pH shift" that denatures the proteins in the algae.

@gjcarew I might have misinterpreted what was written, but when I read that section my brain interpreted it as (based on the rest of the paper) by supplying in reverse==> giving plants O2 during the photoperiod and CO2 during the "rest" period, they achieved a similar result as when a dietician and physical trainer are working together to help a performance athlete achieve desired results. To "break" a plateau, either caloric intake has to be changed (in the plants, this was the CO2), stress load has to be changed (photoperiod), or in many genotypes of individuals it needs to be a combination. By denying the plants CO2 during the photoperiod, the plants attempted to compensate by building more chlorophylls (equivalent of muscle in humans).

I suspect the side effect of denying light during the CO2 saturation period meant the plants could not offgas O2, which would have sped up the process of asphyxiation in all the pests.

For the algae, which does not have the same layers of cellular mass protecting the nucleus of the cell as the multicelled, multi-layers of cells plants, the algae wouldn't be able to withstand the rigidity (change in torpor) due to the pressure of the CO2.

But, perhaps I read it wrong....

On 6/16/2022 at 4:36 PM, dasaltemelosguy said:

One really cool aspect of the whole thing was frankly a surprise was when the plant was moved out of the seltzer to plain water, the pH shock literally shattered the snail eggs and killed the algae. We were planning on incubating them to watch for hatching after seltzer, but they literally exploded upon removal from seltzer and being placed in water. 

This actually makes perfect sense, after the fact, especially if differences in osmotic pressure are accounted for, combined with no light + CO2 in the plants = no offgassing of O2 by the plants. The CO2 is going to have a molecular density of 44 grams per molecule versus the oxygen (which is staying absent as photosynthesis is not taking place due to lack of light) which only has a molecular weight of 32 grams per molecule. Therefore, as soon as the CO2 saturated snail egg is dropped into the lower density aerated water, the difference in osmotic pressure would explode the egg. Just like putting a cold egg straight from the fridge into boiling water will explode.

Wouldn't be surprised if this plays a role in the cellular lysing you saw in the algae.

[gjcarew]

On 6/16/2022 at 3:47 PM, dasaltemelosguy said:

My writing made that confusing, I'm sorry. I actually should have added the flame tests we made form evaporating the solutions that the plants were cleaned in. In brief, we saw simple, chemical reactions from the intense electron field of the carbonic acid. We saw (I may have the % wrong, it's somewhere in my notes) something akin to 40% more denaturing in the dark. So, what you said my very well occur at least partially in the light. What we found in the evaporates was largely carbonate salts from the algae Magnesium, some Sodium and some Potassium. I left a lot out as the piece became so long! But it's all available in the downloads in PDFs at the end. 

I don't know though. That would be extremely cool though! It was performed as a full, factorial so it's more or less ready for such. 

I don't suppose you measured pH after treatment in the dark vs. treatment in the light? That way you could tell if photosynthesis (and subsequent glucose synthesis during the Calvin cycle, which is where the carbon actually gets used) was reducing the acidity of the solution at a faster rate than you would see with only atmospheric equilibration.

It would make sense that higher CO2 levels during the duration of the "dark" treatment leads to more carbonic acid in solution, and greater denaturing of proteins. The expected result would be more carbonate salts in the evaporates since there would hypothetically be more of both reagents in the solution.

 

 

[dasaltemelosguy]

On 6/16/2022 at 3:57 PM, Torrey said:

This actually makes perfect sense, after the fact, especially if differences in osmotic pressure are accounted for, combined with no light + CO2 in the plants = no offgassing of O2 by the plants. The CO2 is going to have a molecular density of 44 grams per molecule versus the oxygen (which is staying absent as photosynthesis is not taking place due to lack of light) which only has a molecular weight of 32 grams per molecule. Therefore, as soon as the CO2 saturated snail egg is dropped into the lower density aerated water, the difference in osmotic pressure would explode the egg. Just like putting a cold egg straight from the fridge into boiling water will explode.

Wouldn't be surprised if this plays a role in the cellular lysing you saw in the algae.

You understand it perfectly. Where were you 6 months ago! 🤣

[Ken Burke]

On 6/16/2022 at 6:27 PM, gjcarew said:

I don't suppose you measured pH after treatment in the dark vs. treatment in the light? That way you could tell if photosynthesis (and subsequent glucose synthesis during the Calvin cycle, which is where the carbon actually gets used) was reducing the acidity of the solution at a faster rate than you would see with only atmospheric equilibration.

It would make sense that higher CO2 levels during the duration of the "dark" treatment leads to more carbonic acid in solution, and greater denaturing of proteins. The expected result would be more carbonate salts in the evaporates since there would hypothetically be more of both reagents in the solution.

 

 

That’s exactly what I was just telling my dog!  Well maybe my dog  was telling me.  I can’t remember.  Glucose, that’s brown sugar, right?😳

[Griznatch]

Wow, very impressive. I just wanted to say thanks for doing this!

 

I'm still reading through this by the way ...

Edited June 17, 2022 by Griznatch
add more

[Its Hutch]

WOW! The amount of work and effort you all put in is huge. Congratulations. I wish this would have came out a week ago before I totally destroyed some anubias leaves trying to kill some algae but I will definitely be trying this in the future very soon. Thank you for putting in the time and effort. This is above and beyond but exactly what this forum can do. Thank you. 

[Guppysnail]

@Its Hutch I hope this inspires others to team up for some projects. It really makes what is possible with this forum fun and just “more” than what singular projects can be. 

[Ken Burke]

e

so giving this a whorl. I selected this plant because it was small enough to fit in a plastic cup, and it is already weighted.  I have a filter on the faucet at my coffee bar, but used a test strip to make sure I did not have any chlorine in the water.

I filled the soda stream with the filtered water and charged it.  Soda stream uses a pressure release valve to prevent over pressurizing the carbonated water.

 

water in the cup, and added plant.  I put it in a darkroom (0 light)at 1415 ish. We’ll see the results in a while.  
 

Question:  have we tried it after only 4 hours vs the 12 hours in the write-up?  I have other plants I can test with, wonder if results require 12 hours in the dark?

[Guppysnail]

On 6/17/2022 at 5:04 PM, Ken Burke said:

e

so giving this a whorl. I selected this plant because it was small enough to fit in a plastic cup, and it is already weighted.  I have a filter on the faucet at my coffee bar, but used a test strip to make sure I did not have any chlorine in the water.

I filled the soda stream with the filtered water and charged it.  Soda stream uses a pressure release valve to prevent over pressurizing the carbonated water.

 

water in the cup, and added plant.  I put it in a darkroom (0 light)at 1415 ish. We’ll see the results in a while.  
 

Question:  have we tried it after only 4 hours vs the 12 hours in the write-up?  I have other plants I can test with, wonder if results require 12 hours in the dark?

That is a fantastic question. 
4 hours started to denature. But that algae recovered. 5 hours killed some types but not 100% reliably. The reliability increased to having found 9 hours is minimum for 100% reliability on all types I tested. 
I also placed several plants that were polluted with BBA in a tank that had never experienced BBA after they were treated to be sure no spores survived. I’m at the 2 1/2 month mark of 9 hour treated plants in that tank and there is zero BBA that I observed anywhere in the tank. 
Thanks for asking as always your questions @Ken Burkeare always thoughtful. 

To add I also placed 9 hour treated plants back in tanks with BBA and those specific plants did not experience BBA again at the 4 month mark. 

[Ken Burke]

And so….

because I started in the late afternoon, my soak was about 18 hours.   Eeek

the ph was definitely lower.

. 
 

and the algae was turning red as soon as it plant hit the water.  I’ll try to get a picture in another hour or so if there is a difference from this one.

[Guppysnail]

@Ken Burke No worries on the 18 hour one. My hubby and I got delayed on errands one day and I forgot to take a plant out of the soak until the next morning.  Thank you again for trusting us enough to try this.  
 

I hope you share the progress of this anubias following any “ph pruning” and continued growth etc. I would love to see this thread become a fully interactive thread where everyone shares their experiences with Reverse Respiration on both the pest removal and algae death as well as after plant progress  That would truly make this a full Forum Collaboration 🤗 We obviously could never test every plant or algae grown under every condition.

I tested a total of 16 anubias plants with different types of algae. Neglecting maintenance on tanks to encourage algae growth so I did not run out and grew as many types as I could, all my slow growing anubias and Java fern were obviously hit. Here is my experience with those through the “ph pruning” process. 
 

Looking at the vascular activity of the leaves before treatment I distinctly noticed the most infested leaves had the least to no internal activity under the microscope.  Over roughly 2-3 weeks those leaves would yellow starting around the tips and edges, only a few became yellow throughout all at once, and disintegrated. New leaf growth would begin.  On my oldest “mother anubias plants” that regularly flower and produce entire “daughter” plants occasionally each of those 3 produced new leaves and a daughter plant within 1-2 months. 
 

In the early stages of this trial I thought I had caused long term damage seeing the yellowing and that is why I started looking at the leaves before and after under a scope. This lead to being able to realize that “ph pruning” was occurring on leaves already starting to die.  
 

Thank you again and I really hope EVERYONE shares whatever they experience through this process.

[modified lung]

On 6/16/2022 at 3:31 PM, gjcarew said:

Tell me if I'm understanding correctly: You're claiming that in the absence of light, chlorophylls are "attempting" photosynthesis but somehow oxidizing the surrounding material? What is the evidence? I'm not questioning the phenomenon of algae dying, but that seems like an unlikely explanation. It seems more plausible that given high co2 + light, algae can consume enough CO2 through photosynthesis to create local microclimates of higher pH than in the surrounding solution. This might delay or even stop the "pH shift" that denatures the proteins in the algae.

I wasn't going to bring it up but I also think the "everything is energy" explanation seems like a big leap. This sounds to me like something that could happen in theory but, if possible, only be observed under a massive input of energy. Like hadron collider type stuff.

But pH microclimates happen in water all the time. For example, if oxygen consumption is high enough, fish can push out enough CO2 to create a microclimate of low pH around their gills protecting them from ammonia burn for a time.

Then the algaecide effect might be partly due to a lower tolerance to osmotic pressure changes. A severe osmotic pressure change wouldn't just cause electron loss, it can rip basically everything out of the cell. I've even seen fish skin melt away after a rapid osmotic pressure change.

Then perhaps the plants are reallocating stored energy (maybe from the roots?) to build more chloroplasts in an attempt to compensate for no light + high CO2 conditions as a survival mechanism. Basically, they could be sensing the CO2 availability and be trying to grab onto any possible light they can to sustain themselves until conditions return to normal? Then when put back into a normal environment, there might be a plateau breaking effect like Torrey described. 

 

I thought everything else was really great though. It makes me want to step up the game on some of my post (...lol I won't though).

 

 

[Guppysnail]

@modified lung thank you for taking the time to bring this up.  Input like yours will help us refine the article to make it a better read and clearer.   We were actually just speaking of how what was being explained could have been explained better. As soon as he gets the opportunity @dasaltemelosguy is going to post a more detailed explanation. 
I am not the scientist of the two so I do not want to try and elaborate (I would definitely make a mess of it). 
 

Thank you again and I really enjoy your your posts. I think your game is already “stepped up” 😁