EDTA refuses to dissolve!!! - argh!! (Jul/22/2009 )

one more thing. you start with 800ml water, add 186.1gm edta and then about 20gm naoh. the final adjustment to 1 liter should be minimal.

you neither make adjusted solutions at the final volume (before adjustment) nor do you start with the final volume and then add to it.

Sambrook et al wrote their protocols because they worked, so it's not wise to decide they are wrong and just do your own thing before trying things their way. EDTA has very low solubility at low to neutral pH, which is why you need to throw in large amounts of NaOH to get it to dissolve. Remember it is an acid, so you shouldn't have to worry about adding extra acid.
Why are you wanting 1M EDTA? Most of the time, you only need 1 - 5 mM, so a 1M stock is not really necessary.

bochum on Jul 22 2009, 10:49 PM said:

Hi bochum,

Yes, it is not possible to prepare 1M EDTA, but I did not know that prior to doing it until I read the sambrook molecular cloning book.

What I did was to add 186.1g and added 800mL of water. What I am trying to tell you is that after adding NaOH to adjust the pH to 8.0, it was right at 1L. So there was nothing wrong with what I was doing. I still get 0.5M EDTA stock because 186.1g of EDTA was dissolved in a final volume of 1L. It took quite a lot of NaOH before the solution adjusted to pH8.0

mdfenko on Jul 23 2009, 05:23 AM said:

Dear mdfenko,

That was what I did.

In my previous entry, I stated
"I went back to the lab just a while ago to follow your suggestions of adding the NaOH slowly. I also increased the volume to a little less than 1L with dH20 so I can still add NaOH to adjust the pH at the same time. It is horrifying how much NaOH I had to add to adjust the pH. It took forever but at least it is dissolving now. I am letting it stir overnight."

I left my solution of 186.1g EDTA and 800mL water in the lab while I posted here for help, after reading a few replies, I went back to the lab again and added the NaOH slowly. Water was also added because to get 0.5M, it is necessary to hit a total of 1L.

The reason why I added water was because pH8.0 was hit and the EDTA started to dissolve and I have to add water to hit it to 1L. As it dissolved, the pH went down again due to the reaction and I added NaOH to adjust the pH to 8.0 again before everything dissolved.

swanny on Jul 23 2009, 09:31 AM said:

Dear swanny,

THanks for your comment.

Well... I made 1M EDTA initially because I wanted to have a stock solution and it was my first time making an EDTA stock solution. So naturally.. I did not know that 1M EDTA was impossible.

jiajia1987 on Jul 22 2009, 09:48 PM said:

but you shouldn't adjust the volume until all of the edta is in solution so that you still have room to add naoh.

mdfenko on Jul 25 2009, 06:02 AM said:

hmmm.... ok, point noted!

bochum on Wed Jul 22 14:49:29 2009 said:

Hi If i am right, it is not possible to prepare 1M EDTA,ie what even sambrook and russell's molecular cloning book suggest to prepare 0.5M EDTA. And regarding calculation when the molecular weight is 372.24g/mol,then to prepare 1M solution you have to take 372.24g in 1litre and to prepare 0.5M you have to take 186.1g ie what sambrook and russell's molecular cloning book suggesting you.

But if you prepare it by adding excess of NaOH or HCL by any means it is wrong...and it has to go directly to dust bin

So point is you cannot or should not prepare 1M EDTA and there is nothing wrong in the book...

 

I added excess NaOH in preparing the 0.5 M EDTA overshooting the pH to 11.0, so I adjusted the pH using concd HCl. Unfortunately, I added excess HCl and EDTA started to solidify. This made me add NaOH again and finally I adjusted the pH of the solution to 8.0. Is this solution still valid for molecular biology use? If it is not, pls explain why..Thanks in advance.