I want to check the effect of melatonin on RAW264.7 cells.I have used ethanol 10 microliter/mL, but this ethanol has some effect and I found the effect of melatonin very few or nothing. So, what solvent and in which concentration will be good for my experiments? Do I need to make molar solution of melatonin or 10, 1, 0.1, 0.01 microgram/mL is ok? Please suggest me.
Melatonin Solubility :
in water 0.1 mg/ ml
in Ethanol 8 mg/ ml
otherwise u can make a 50mM stock solution on DMSO
Melatonin Solubility :
in water 0.1 mg/ ml
in Ethanol 8 mg/ ml
otherwise u can make a 50mM stock solution on DMSO
Oh! Thanks a lot for your answer. How much ethanol or DMSO is good for RAW264.7 cells? I mean solvent will have no effect on osteoclastogenesis.
final DMSOO/ Ethanol concentration in a disc contain should not exceeds 2%
We disolve melatonin in ethanol first and then ad water or saline. You will get quite stable solution/suspension.
I tryed to dissolve up to 20 mg melatonin in 0,5 ml ethanol and got pure solution.
Oh! Thanks a lot to both of you. I used 1% in alpha MEM, not in disc @ Jhuma Bhadra. It lowered osteoclasogenesis significantly.
Which Saline do you use @ Andrey V Panchenko? Do you use 20mg melatonin to dissolve in 0.5mL ethanol? Then you mix it with water or saline to dilute? Can I use alpha MEM insead of water or saline? And another thing, how and how long I can store this melatonin solution?
Thanks for your helping hands.
I have to apologize for misunderstanding. We prepare such solution of melatonin for our animal experiments. We do not use any stock, we use fresh solution always because melatonin is light sensitive and we are not sure about solution stability.
May be next information could be helpful:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3887041/pdf/pone.0085311.pdf
Vehicle solution was prepared with 8 ml of phosphate buffered saline (PBS), 1 ml of dimethyl sulfoxide (DMSO) and 1 ml of Cremophor (Sigma, St. Louise, MO, USA).
Melatonin (Sigma, St. Louise, MO, USA) was diluted in vehicle and the animals from the melatonin group received IP of 100 ml of melatonin treatment (at dose of 40 mg/kg of body weight) for five days a week.
Oh! Thank you very much. I will try. Thanks again for your helping hands to me.
The Molecular weight of melatonin is 232.28. Solubility of melatonin in DMSO is 50 mM. That means if you dissolve 11.61 mg in 1 ml of DMSO it will give you a 50mM solution. So the standard range of exposure for vehicle (DMSO) is 0.1%. This means if you add 1 ul in 1ml that will give you a 0.1% solution. So if you add 1 ul of 50 mM solution in 1 ml of DMEM, the final strength of the solution will be 50 uM. This can be your starting point. In case you don't see an effect at 50 uM exposure. You can increase the vehicle percentage upto 0.5% but not higher than that. In my experience RAW264.7 cells are fully viable and healthy at 0.5% exposure to DMSO. At 0.5% you will get a working solution of 250 uM. Don't forget to expose the cells with vehicle only and vehicle + RANKL as negative and positive control.
You could use etanol or DMSO to dissolve melatonin, and then in culture medium for the cell treatments. Final ethanol or DMSO should not ust exceed 0.5-2% in the culture medium.
It is possible to solve melatonin in one drop of ethanol and than make consequence solutions of concentrations you need
I never worked with cell cultures. In in vivo studies, I solve melatonin in couple drops of ethanol and then prepared concentration I need using tap water for administrtion of melatonin with drinking water or, using saline if it will injected.
Hi there
what is the source of melatonin you apply? is pharmacological melatonin applicable?
Hi
Can you please tell me the best vehicle for melatonin powder to be used in animals as a long acting subcutanouse implant?
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