I measured the absorbance of CBD (1mg/ml in methanol) diluted in HEPES buffer (0.2M NaCl and 20mM Hepes pH 7.5). I have attached a picture of my results - however it does not resemble what is found in literature (CBD exhibits two absorption maxima in the 210–220 and 270–280 nm regions and emission in the 290–300 nm region). Any help towards analyzing these results will be appreciated!
According to the product literature from Sigma for a ~0.1 mg/mL (about 0.3 mM) solution of cannabidiol in methanol, there is an absorbance peak at 273 nm with an absorbance of about 0.4. The product page (https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/266/034/c6395-slbk4474vdat.pdf)
doesn't say what the path length was, but I'll assume it is 1 cm, which is a pretty standard path length for a quartz cuvette. What this means is that your solutions are probably too dilute to detect the 273 nm absorbance on the scale you show. A 10 µM solution would have an absorbance of about 0.013, if it were in methanol.
You diluted the CBD solution from methanol to HEPES, which will have an effect on the absorbance spectrum, and may be a problem for solubility of the compound as well if you try to make a more concentrated solution.
Hello!
First of all, anything above 1 au can start to swamp your detector, and you can see that with the noise at your highest concentration.
Secondly, your cuvette, solvent, and buffer can absorb too, so be sure to check that you aren't just seeing swamping out from your cuvette/sample/buffer. Take a blank with just water, then do with your buffer/methanol mix and see if it absorbs where you are looking for CBD.
Further, are you seeing solubility issues in the buffer like phase separation (CBD is fairly non-polar so I wouldn't expect it to be water soluble)? That'll mess up any linearity and peak shapes as well.
Cheers,
Kyle
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