I have the same problem as you described below. Did you descover the reason. I could uspect pH, but i am not sure. Hope you can help.
I have prepared the mixture as shawn below.
A) Dissolve 50g of acid free EDTA in 250 ml of ddH2O. Heat to dissolve.
B) Dissolve the following, one by one, in order, heating to approximately 100° in 550 ml ddH2O.
BO3H3 11.40 g
ZnSO4·7H2O 22.00 g
MnCl2·4H2O 5.06 g
FeSO4·7H2O 4.99 g
CoCl2·6H2O 1.61 g
CuSO4·5H2O 1.57 g
(Ammonium Molybdate) 1.1 g
Mix the two solutions (A & B) together. The resulting solution should be blue-green.
Heat to 100°C. Cool slightly, but don't let the temperature drop below 80°-90°.
Adjust pH to 6.5 to 6.8 with 20% KOH (approximately 83 ml). Don't let the temperature drop below 70 ° until after the pH is adjusted.
Make up to 1L and let stand in a 2L Erlenmeyer flask, stoppered loosly. The color should change from green to purple over a period of days.
Remove the rust-coloured precipitate by filtering, with suction, through 3 layers of Whatman #1 filter paper in a Buchner funnel. Repeat until no more precipitate is seen on the filter paper
Store in a brown bottle at 4°.
I hope some among you have experience in preparing Hutner trace elements
solution. I followed the procedure described on page 579 in the Chlamydomonas
The problem occurs during the final step, when I adjust to pH 6.7 by adding
20% w/v KOH. The solution changes from clear dark green to a suspension of a
green solution and rust-brown particles. According to the procedure it should
be clear green initially and after one or two weeks the color changes to
purple leaving this rust-brown precipitate. However, when I let the solution
stand for 2 weeks nothing changes and after two weeks I still have a
suspension of a green solution and a rust-brown precipitate...
I already tried this procedure for three times and always I have the same
Does anybody know what could be the cause of this behaviour,
thank you very much for your time,
Food and Bioprocess Engineering Group
Wageningen Agricultural Universtity