I extracted plant material with ethanol for anticonvulsant activity. DMSO is used to dissolve the ethanolic extract. But ethanol itself has CNS stimulant activity. So please suggest a method by which we can calculate the amount of ethanol present in the ethanolic extract to confirm the convulsant activity of the plant extract.
You can calculate it by HPLC analysis of sample Ref: http://www.phenomenex.com/Application/Detail/5513?returnURL=/Compound?id=Qcbppad]H%40&alias=Qcbppad]H%40
I would use alcohol dehydrogenase. As the alcohol gets oxidized by the enzyme, the NADP gets reduced. Follow the absorbance change at A340 nM. You can make a standard curve with known amounts of ethanol. Use this if you don't have a mass spec or use instead of HPLC.
better way to compare efficacy of pure drugs in the solvent by using control group with equal amount of DMSO and ethanol for same efficacy while blank containing absent of both solvent and drugs too.
Dont go through estimation of solvent or practice to remove same.
Why you are using DMSO as solvent. Keep ethanol for same for dilution or use buffer or distill water for serial dilutions.
I use methanol to extract plant material and then lyophilise the extracts such that there is now no methanol in it. so i would suggest you do the same.
I am completely agree with Warren John. We work with ethanol extracts and remove the solvent before the investigation of metabolic effects
HPLC ? We use that to quantify residual solvents in radiopharmaceuticals. That may be too expensive or overkill for your purposes, but if you already have HPLC and someone familiar with it's operation, it's an alternative worth consideration.
About chemistry: Once you "rotavap" your extract, it always may get enriched with -or retain-some- water. It is true that the vacuum in a freeze dryer may help to evaporate some of these solvents but freeze drying is not intended to remove ethanol or methanol or DMSO, just water only. The loss on drying is the simplest one to quantify "Total solvent content" although it does not give you the identity or proportion of each one. This has to be quantified by GC (FID would suffice) to get the most accurate analysis. A common way to totally dry any traces of solvents is "rotavapping" a small amount of extract in a small vial to get a thin film of the extract evenly distributed on the walls, then we place the vial for an hour under constant stream of DRY PURE Nitrogen gas.
About Pharmacology: virtually any organic solvent (and water sometimes, as it dilutes your buffers and its conc and osmolarity changes) will have a pharmacological effect on your assay. The only way to minimise it is running controls/blanks with the solvent only and correcting the "background effect/noise". DMSO is not necessarily better than ethanol as it exerts a plethora of differentiation and redox effects in intact cells. Ideally it would be better to dissolve the extract in the cell media (did you try ?) sometimes the presence of Serum helps to dissolve organic extracts. For aqueous extracts we many times find possible to redissolve them in the very same cell media (DMEM, etc.), so we minimise the impact.
Hope it helps!
I agree with Jose . You can use HS analysis along with GC FID to know % of solvent analysis.
- Badges
- Science topic
- Similar topics
- Biological Science
- Botany
- Pharmaceutical Botany
- Plant Extracts