A pressure cooker would only serve to raise the temperature. I think the gentle agitation of boiling is bigger factor (than a few degrees more heat), so the pressure cooker may actually be counter productive. I'm not sure of the hazards involved with pressure cooking H2O2 either (if any). Stronger H2O2 concentrations are shipped in vented bottles for a reason...
The clumps issue applies to fossil and subfossil - including mudstones.
Cleaning diatoms
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Thank you very much! That makes sense and I will try your procedures.Beatsy wrote:A pressure cooker would only serve to raise the temperature. I think the gentle agitation of boiling is bigger factor (than a few degrees more heat), so the pressure cooker may actually be counter productive. I'm not sure of the hazards involved with pressure cooking H2O2 either (if any). Stronger H2O2 concentrations are shipped in vented bottles for a reason...
The clumps issue applies to fossil and subfossil - including mudstones.
Selling my Canon FD 200mm F/2.8 lens
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Smokedaddy
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I'm not chemistry savvy in the slightest but definitely safety conscious when dealing with stuff like this. I worked in the semiconductor industry for many years but only at the tool and support piping design level. Safety is taken extremely serious as are chemical spills like hydrofluoric acid (which can get you banned from the project as well as many other safety issues). I suppose hearing the word acid kicks in that programmed fear in me and the lack of a chemistry understanding when it comes to processing diatoms. <g>
JW,Smokedaddy wrote:I'm not chemistry savvy in the slightest but definitely safety conscious when dealing with stuff like this. I worked in the semiconductor industry for many years but only at the tool and support piping design level. Safety is taken extremely serious as are chemical spills like hydrofluoric acid (which can get you banned from the project as well as many other safety issues). I suppose hearing the word acid kicks in that programmed fear in me and the lack of a chemistry understanding when it comes to processing diatoms. <g>
Beatsy's fresh sample protocol (the water boil one) does not require acid. Just remember to turn off stove.
Hydrofluoric acid is a highly corrosive poison, while hydrochloric acid at 31% is a bit safer. Your concern is valid though.
Selling my Canon FD 200mm F/2.8 lens
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Smokedaddy
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Don't mean to sound stupid but I was pretty excited today. I took a water sample from a creek in the Bradshaw Mountains while I was 4 wheeling a couple weeks ago. Today I took one drop from the bottom of the jar, put it on a slide along with a coverslip. Man was I blown away. My first diatom (or I should say diatoms). To bad there's so much crud in the sample. Now if I could only process them properly and safely.
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Smokedaddy
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I was reading this ...
Throughout the text, the phrase "discard the supernatant" will occur. Do not use a pipette for this, or do not pour off the supernatant, as either action may disturb the sediment and you may lose material. The best way is siphoning with small diameter soft plastic tubing. The speed of draining can be sensitively controlled by pinching the tube.
Out of curiosity how is siphoning done? Are they talking about using something like a straw, dip it in the supernatant, put your thumb on the end etc.?
-JW:
Throughout the text, the phrase "discard the supernatant" will occur. Do not use a pipette for this, or do not pour off the supernatant, as either action may disturb the sediment and you may lose material. The best way is siphoning with small diameter soft plastic tubing. The speed of draining can be sensitively controlled by pinching the tube.
Out of curiosity how is siphoning done? Are they talking about using something like a straw, dip it in the supernatant, put your thumb on the end etc.?
-JW:
It is not absolute. I discard supernatant and use pipette myself and I can tolerate losing some diatoms. You won't lose a lot if you are careful.Smokedaddy wrote:I was reading this ...
Throughout the text, the phrase "discard the supernatant" will occur. Do not use a pipette for this, or do not pour off the supernatant, as either action may disturb the sediment and you may lose material. The best way is siphoning with small diameter soft plastic tubing. The speed of draining can be sensitively controlled by pinching the tube.
Out of curiosity how is siphoning done? Are they talking about using something like a straw, dip it in the supernatant, put your thumb on the end etc.?
-JW:
Yes, that is how Rod does his siphoning. That is a better way than pipetting. Rod does it because he cannot sleep well, if he finds he loses some diatoms
Selling my Canon FD 200mm F/2.8 lens
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Smokedaddy
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JW,
You would have to ask Beatsy for his exact protocol. I don't know the specifics of his protocol.
I am not familiar with the tetrasodium pyrophosphate component of his protocol. Especially, how to remove it and its complexes in the final step.
I am guessing (I could be wrong) tetrasodium pyrophosphate is used there to precipitate calcium, but then separating calcium precipitates from diatoms may not be easy (though lots of distilled water wash may do it - not sure). I also don't have tetrasodium pyrophosphate at hand to experiment.
If Beatsy does not reply to you, I can modify my previous protocol for you to use with his chemicals. I am guessing he is also using 30% hydrogen peroxide (which is one of the chemicals used to make DIY bombs and not a very safe chemical either - though it is not that dangerous by itself). Also note I don't know how to use tetrasodium pyrophosphate exactly.
Overall, Beatsy's protocol has the least amount of toxic chemicals. That is for sure. The sodium dichromate or muriatic acid in my protocol are both toxins - you need yo wear glove while handing them.
You would have to ask Beatsy for his exact protocol. I don't know the specifics of his protocol.
I am not familiar with the tetrasodium pyrophosphate component of his protocol. Especially, how to remove it and its complexes in the final step.
I am guessing (I could be wrong) tetrasodium pyrophosphate is used there to precipitate calcium, but then separating calcium precipitates from diatoms may not be easy (though lots of distilled water wash may do it - not sure). I also don't have tetrasodium pyrophosphate at hand to experiment.
If Beatsy does not reply to you, I can modify my previous protocol for you to use with his chemicals. I am guessing he is also using 30% hydrogen peroxide (which is one of the chemicals used to make DIY bombs and not a very safe chemical either - though it is not that dangerous by itself). Also note I don't know how to use tetrasodium pyrophosphate exactly.
Overall, Beatsy's protocol has the least amount of toxic chemicals. That is for sure. The sodium dichromate or muriatic acid in my protocol are both toxins - you need yo wear glove while handing them.
Last edited by zzffnn on Wed May 10, 2017 4:44 am, edited 1 time in total.
Selling my Canon FD 200mm F/2.8 lens
The peroxide/tspp boil is very simple. Note: I can't get 30% peroxide any more either (applying for a personal permit for the "difficult" chemicals soon) but 12% will do for this.
A full protocol would be something like...
1. Freeze/thaw to break up the sample thoroughly (if mudstone)
2. Soak in hydrochloric acid to remove calcium if necessary
3. Rinse sample to neutral. This involves topping up with DI water, allow to settle, then decant off the supernatant. Repeat several times until liquid tests neutral pH (cheap litmus paper is fine). In future steps all this will just be referred to as "rinse"
4. Put 1L of 50/50 DI water and peroxide in a suitable container for boiling (I use a glass milk pan, but a large flask will work, beware of bumping though). 12% peroxide will do - but you will need to boil for longer
5. Add a generous pinch of TSPP - 1g or 2g should be enough - I haven't determined an optimum amount yet. It's easy to get (Ebay) and is used in the food industry to keep stuff in suspension (chicken nuggets etc). It is also used by geologists to break up shales and the like.
6. Add your sample
7. Bring to a slow rolling boil and keep it there for 2-6 hours. Top up with DI water as the liquid evaporates.
8. Pipette off a bit of the liquid while boiling, allow to settle in a test tube and examine the settled residue to see how cleaning is progressing. Stop when the frustules look clean.
9. If the frustules have stuff stuck to them after a few hours, add a few grams of lux soap flakes or similar. Do not boil for more than an hour with this though as it is alkali and can damage the frustules.
10. Cool - then rinse and settle several times
11. I now wash the whole sample over a 20um mesh to get rid of fine silt. Of course, this will lose most diatoms <20um. If you want to keep those just do a lot more rinses - discarding the cloudy part of the water after a suitable settling time.
12. Now you should have a cleaned sample with no silt, but it will contain grit, sand, and (of course) your diatoms.
13. Separate the sand/grit by putting 1cm of DI water in a flask, add (some of) the sample and gently swirl it around until a small pile of heavy material forms in the middle. Pipette this little pile off (save in another container) and continue swirling and pipetting until nothing else gathers in the centre. Once you reach this stage, you should pretty much have nothing but frustules left suspended in the water. Repeat the whole process several times with the "heavy stuff" that is pipetted off (examine under the microscope to see if it still contains frustules) to ensure you get as many of the frustules out as possible.
14. If this doesn't leave you with perfectly clean frustules, then a shock in sodium hydroxide may be needed (see previous post).
15. If all this doesn't work - your sample needs to be acid cleaned instead.
That's about it.
Hope that helps.
A full protocol would be something like...
1. Freeze/thaw to break up the sample thoroughly (if mudstone)
2. Soak in hydrochloric acid to remove calcium if necessary
3. Rinse sample to neutral. This involves topping up with DI water, allow to settle, then decant off the supernatant. Repeat several times until liquid tests neutral pH (cheap litmus paper is fine). In future steps all this will just be referred to as "rinse"
4. Put 1L of 50/50 DI water and peroxide in a suitable container for boiling (I use a glass milk pan, but a large flask will work, beware of bumping though). 12% peroxide will do - but you will need to boil for longer
5. Add a generous pinch of TSPP - 1g or 2g should be enough - I haven't determined an optimum amount yet. It's easy to get (Ebay) and is used in the food industry to keep stuff in suspension (chicken nuggets etc). It is also used by geologists to break up shales and the like.
6. Add your sample
7. Bring to a slow rolling boil and keep it there for 2-6 hours. Top up with DI water as the liquid evaporates.
8. Pipette off a bit of the liquid while boiling, allow to settle in a test tube and examine the settled residue to see how cleaning is progressing. Stop when the frustules look clean.
9. If the frustules have stuff stuck to them after a few hours, add a few grams of lux soap flakes or similar. Do not boil for more than an hour with this though as it is alkali and can damage the frustules.
10. Cool - then rinse and settle several times
11. I now wash the whole sample over a 20um mesh to get rid of fine silt. Of course, this will lose most diatoms <20um. If you want to keep those just do a lot more rinses - discarding the cloudy part of the water after a suitable settling time.
12. Now you should have a cleaned sample with no silt, but it will contain grit, sand, and (of course) your diatoms.
13. Separate the sand/grit by putting 1cm of DI water in a flask, add (some of) the sample and gently swirl it around until a small pile of heavy material forms in the middle. Pipette this little pile off (save in another container) and continue swirling and pipetting until nothing else gathers in the centre. Once you reach this stage, you should pretty much have nothing but frustules left suspended in the water. Repeat the whole process several times with the "heavy stuff" that is pipetted off (examine under the microscope to see if it still contains frustules) to ensure you get as many of the frustules out as possible.
14. If this doesn't leave you with perfectly clean frustules, then a shock in sodium hydroxide may be needed (see previous post).
15. If all this doesn't work - your sample needs to be acid cleaned instead.
That's about it.
Hope that helps.
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Smokedaddy
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