Difficulties extracting THC from "energy shot" type product

Discussions about sample preparation: extraction, cleanup, derivatization, etc.

15 posts Page 1 of 1
I am looking for a method to extract THC out of a "energy shot" type of product to analyse via GC-FID(HP 5890).
The product is being legally sold in washington under I-502 guidelines. I've tried a few liquid-liquid methods, but am not recovering anywhere close to what I should be. When testing plant materials I get excellent recovery, but something in this matrix is causing me problems.

Using hexane as my solvent I am recovering ~ 8ug/ml
The manufacture claims that the thc content is 5mg/30 ml or 0.166mg/ml
The ingredients are below
My last approach was as follows

1/2 ml Sx
1/2 ml hexane
vortex
centrifuge
add NaCl
vortex
centrifuge
filter through .45 syringe filter
manually inject into gc

The chromatogram looks nice. Nice background and clean peak.

Proprietary Ingredient Blend: Organic Cane Alcohol, Cannabis, Guarana Seed Powder, Organic Fair Trade Yerbe Matte Leaf, Quercetin from Green Tea

Other Ingredients: Water, Honey, Xylitol , Organic Lemon Extract, Citric Acid, Other Natural Flavors
UCT (www,unitedchem.com) has an excellent SPE app note for THC. Dilute the sample and follow the instructions. It should provide a good separation. Call their tech support with questions. They are very skilled at separating THC from other compounds for analysis.
Don Shelly
LGC Standards
dlntx9 wrote:
Using hexane as my solvent I am recovering ~ 8ug/ml
The manufacture claims that the thc content is 5mg/30 ml or 0.166mg/ml


8µg/ml v. claimed 166µg/ml amount, that's large! If me, I'd spike a sample with known 166µg/ml amount, and see what I get/recover, almost like a standard additions investigation. Then you'd know that at least 166µg/ml should be present.
Don,

Thanks for the UCT suggestion. Looking over the website now.

I've tried spiking the sample and am not getting a good recovery. Something must be binding with the THC, and keeping it out of the hexane layer.
dlntx9 wrote:
Don,

Thanks for the UCT suggestion. Looking over the website now.

I've tried spiking the sample and am not getting a good recovery. Something must be binding with the THC, and keeping it out of the hexane layer.


You might need to perform hydrolysis first. Contact Brian Kinsella at bkinsella@unitedchem.com for tips.
Don Shelly
LGC Standards
dlntx9 wrote:
Don,

Thanks for the UCT suggestion. Looking over the website now.

I've tried spiking the sample and am not getting a good recovery. Something must be binding with the THC, and keeping it out of the hexane layer.


As long as the recovery is repeatable you can just correct for it.

Peter
Peter Apps
Peter,

I'm sorry, I'm not following you. What do you mean that I can can correct for it? My GC analysis data is consistent, and the peaks for my controls are what they should be, but I'm failing on actually extracting all of the molecule from the matrix that I'm analyzing.

Thus far this matrix is the only one that has really given me issues. I have methods validated for thc extraction from plant materials via a soxhlet method, and have been using solvent methods for extractions from oils, and other edibles successfully.

The product in question has a low pH, as one would expect for this type of product. The only ingredients that I think could be causing problems is the xylitol, other than it the rest of the ingredients are essentially sugar from honey, caffeine from gaurana, ethanol, citric acid and water.

I've tried using MeohChcl3 which works well with extracting from plant material, but had essentially the same result that I received with the Hexane.

I'm still waiting to hear back from UCT. I couldn't find anything on the site that really addressed the issue im having, maybe I just missed it, but i"m hoping they will have some suggestions for me. It doesn't seem like this should be a very complicated procedure, and it's really humbling that out of all the analysis that I have performed, extracting THC from flavored sugar water is what has stumped me.
Suppose you spike your matrix with 100 units of THC, and get a peak for 10 units. Your recovery is 10%. As long as it is always 10%, when you analyse a sample you multiply the peak area by 10 to correct for the 10% recovery.

Peter
Peter Apps
Below is a UCT method that I found in my notes. The matrix is plant material, but it demonstrates the need for an aggressive extraction. This should be on their website. The key seems to be the 1 hour sonication.

Determination of THC in Marijuana Samples Using Solid Phase
Extraction and LC-MS/MS

UCT Part Numbers:
CSTHC206: Clean Screen® THC, 200mg x 6mL
SLDA100ID21-5UM: Selectra® DA (100mm x 2.1mm, 5 μm) LC Column
SLDAGDC21-5UM: Selectra® DA (10mm x 2.1mm ) Guard Column
SPHPHO7001-5: 0.1M Phosphate Buffer pH 7
August 2013

Procedure
1. Extraction
a) Add 100 mg of Marijuana sample into a clean glass sample tube
b) Add 5 mL of methanol and cap
c) Sonicate for approximately 60 minutes at room temperature
d) Centrifuge for 10 minutes at 3000 rpm
e) Aliquot 500 μL- 1 mL of methanol extract into a clean glass sample tube
f) Add *internal standard and mix.
g) Add 4 mL of 0.1 M phosphate buffer (pH 7) and mix

2. Condition Clean Screen® Extraction Column
a) 1 x 3 mL CH3OH
b) 1 x 3 mL D.I. H2O
c) 1 x 1 mL 0.1 M phosphate buffer (pH 7.0)
NOTE: Aspirate at < 3 inches Hg to prevent sorbent drying.

3. Apply Sample
a) Load at 1 to 2 mL/minute.

4. Wash Column
a) 1 x 3 mL D.I. H2O
b) 1 x 3 mL 100 0.1 M phosphate buffer (pH 7.0).
c) Dry column (5 minutes at > 10 inches Hg).

5. Elute THC
a) 1 x 3 mL hexane/ethyl acetate/ acetic acid (49:49: 2)
b) Collect eluate at 1 to 2 mL / minute.

6. Dry Eluate
a) Evaporate to dryness under nitrogen < 40°C
b) Reconstitute in 100 μL of mobile phase

Instrument Conditions
Column: 100 x 2.1 mm (5 μm) Selectra® DA
Column Temperature: 40ºC
Mobile phase: CH3OH w/ 0.1% Formic acid: DI H2O w/ 0.1% Formic acid (75:25)
Flowrate: 0.5 mL/ minute
Detector: API 4000 MS/MS
MS parameters
Polarity ESI+
Spray voltage V 4500V
Source Temperature 650° C
Curtain Gas 10
Gas 1 40
Gas 2 40
CAD Gas Medium
Dwell Time 150ms
Compound Q1 Q3 Time/
ms
D
Don Shelly
LGC Standards
Have you considered straight dilution? Hard on the column and inlet but don't have extraction issues.

Best regards,

AICMM
I did try a straight dilution, and the chromatogram was a mess. Not something I'd want to subject my column to on a regular basis. I ended up using quechers and had a nice result. Next time I will be implementing an spe extraction via UCT columns and vacuum manifold.
dlntx9 wrote:
I did try a straight dilution, and the chromatogram was a mess. Not something I'd want to subject my column to on a regular basis. I ended up using quechers and had a nice result. Next time I will be implementing an spe extraction via UCT columns and vacuum manifold.


Hi I'm with UCT and just developed a method for cannabinoids (THC, CBD and CBN) in marijuana and infused samples, I would be more than happy sharing you the method if you email me at xwang@unitedchem.com
did you check the ph of the solution (drink/aqueous layer) you are attempting to extract from just to make sure that it is in fact NOT a basified for some reason?

also what does 1/2 ml Sx mean?
A few suggestions.

Try emptying the bottle of energy shot liquid and put some hexane right into the empty. cap and shake. test this. Could be losing some of the THC inside the rim of the bottle, or somewhere in there.

There could be an emulsifying agent binding the THC and not letting it go into solution. Try sonicating (if possible) and heat, centrifuge for longer if your not getting a clean two phases.

Are you using an internal standard from the beginning.? Adding your IS at the beginning and extracting it along with your THC will allow you to monitor your extraction efficiency. It will help you determine if analytes are not going into solution or something else (like the THC being stuck in bottle). If you get poor recovery on your IS you know it is prob an extraction issue. If you get good recovery on IS but low numbers for THC, look at the bottle.

Hope this helps!
Just a thought. Make sure that you have THC and not THCA in your sample. The natural product is THCA. It needs to be decarboxylated after initial extraction.
Don Shelly
LGC Standards
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