Hello,
I am trying to make 0.1 M vitamin C (ascorbic acid) solution (dissolved in ddH2O) and add 250 uL of it to 5 mL agar for a final concentration of 5 mM vitamin C in my experiment. I dissolve ascorbic acid powder (Sigma-Aldrich, product number A4544) in double distilled water and then filter-sterilize (filter size 0.20 um) it for use in my experiment. The solubility for this product, as mentioned in its Specification Sheet, is 50 mg/mL. I make 17.6 mg/mL (0.1 M), vortex it for a long time, the powder seems to completely dissolve, but when I try to pass it through the filter, it gets stuck. I also tried to sonicate for 15 min, tried dissolving it in a warm environment, nothing worked. I changed almost every single thing: water, syringe, doesn’t work.
However, when I try to use a different filter, 0.45 um size, the 0.1 M vit C solution does pass through. I add it to agar plates, I seed the plate with OP50 the next day, and put worms the following day. Around 4 days after adding vit C, crystal-like substances begin to appear both on the surface of and inside the agar, with them being most concentrated at the agar-OP50 border (where OP50 concentration is also highest). The plates that contain just the water but no vit C does not have that problem. Also, vit C containing plates that do not have worms in them still will form the crystals after ~ 4 days.
The worms do not seem to mind the crystals, they just move normally and munch on the food. But did anyone have the same observation/problem when using vitamin C? Could anyone speculate what these crystals might be and why they are forming? Should it affect the phenotype on vit C? It would seem as if the vitamin C did not dissolve properly; but it is difficult to accept that as I vortex the solution for a long time, and crystals appear a few days after adding vit C, and in a vertical plane…
Thank you for your generous help.