I tried to use essential oil limonene to tested for their anti-apoptotic effect on PC12 cell induced by steroid. But when I prepare for the stock solution of limonene I always end up with colloid in the solution. I have try to prepare stock solution 10 mM using limonene 1.7 uL mixing with 998.3 uL DMEM and the result show no anti-apoptotic effect at all. Someone told me that it maybe cause by the limonene and the medium is not mixing homogeneously and I should add some solvent such as DMSO or EtOH. So I try to prepare stock solution of limonene 100 mM using 50 uL DMSO and 8.4 uL of limonene but the limonene and DMSO is separated even I lower the conc. to 50 mM this problem still persist. Then I try to use Absolute EtOH instead of DMSO in the same volume and method now the solution isn't separated but when I adjust the volume using DMEM it become colloid both for 100 mM and 50 mM stock solution.
Should I lower the concentration of stock solution or using any other method to make the stock solution become homogeneously.
Any suggestion would be highly appreciated thank you.
Dear Niphat,
Limonene solubility: Slightly soluble in water (13.8 mg/mL at 25°C); soluble in acetone, dimethyl sulfoxide and ethanol (National Toxicology Program, 1991; National
Library of Medicine, 1998).
Based on the experiments you have done and solubility of limonene as indicated above, I suggest that you make stock solution with less concentration such as 5-10 mM. Both DMSO and ethanol should be good cosolvents to assist in dissolving the limonene in DMEM. You can use sonication for 10 minutes to make sure that all components are in solution.
Hoping this will be helpful,
Rafik
Rafik: That should be mg/L, not mg/mL!, i.e. 13.8 ppm. I usually reckon 15 pm. Close to the solubility limit, dissolution will be very slow.
Naphit: your strategy looks OK, but check that you are not above this solubility limit.
Thank you for the both answers I really appreciated. I will try to lower the stock concentration to 10 mM as you suggest and if it not working I will try 5 mM. Thank again :D
Niphat,
I agree with both, Rafik and Alan. You must prepare your solutión in DMSO or EtOH at 5-10 mM. You must disolve slowly and finally sonicate the solution to be sure that the homogeneity
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