A Comparison of Manual Versus Automated Sample Preparation Methods for Cannabinoid Analysis Roy Turton , Andy Politis , Evan Contreras , Ryan Ravenelle , Kristina Klette 1
1
1
2
2
RM3 Labs, Boulder, CO, USA 2Hamilton Company, Reno, NV, USA
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ABSTRACT
RESULTS
Automated
Weighed subsample to tube
Weighed subsample to tube
Reproducibility Study
Concentrate
Vortex
Add ISTD
Requires Technician Fully Automated Centrifuge
Heat and vortex
1 hr
Add solvent
Filter and collect filtrate
Cap vial
Aliquot filtrate to HPLC vial
HPLC Analysis
Add solvent
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Microlab STARlet ■■ ■■ ■■
% Variation y = 0.9589x + 0.0059 R2 = 0.9474
5%
™
8 Independent Pipetting Channels Tube and Plate Handling Gripper for Labware Movement [MPE]2 Positive Pressure Module
Cap vial
HPLC analysis
15%
1 2 3 4
Day 2 Day 4
10%
5%
10%
15%
20%
25%
30%
0
35%
20
40
% Total THC Manual
20%
% Variation
80% 65% 40%
y = 0.9771x + 0.0319 R2 = 0.9609
20%
80
100
40%
60%
80%
% Total THC Manual
100%
Day 2 Day 4
Day 1 Day 3
15%
10%
0
20
40
Sample #
60
80
100
*% variation =
| manual - automated | manual
37 concentrate samples were run in duplicate via both sample preparation methods
Manual Automated
6%
7.0 1.6 3.8 4.6 Overall = 4.3
Figure 4: Variation in measured total THC between both methods
Sample Duplicate Runs Average Deviation from Mean 8%
manual
Avg. % Variation*
1 2 3 4
0% 20%
| manual - automated |
5%
0% 0%
*% variation =
Day
Variation of Measured Total THC in Concentrate: Manual vs. Automated Sample Prep
Day 2 Day 4
Day 1 Day 3
Sample #
60
Figure 2: Variation in measured total THC between both methods
Actual Measured Total THC in Concentrate: Manual vs. Automated Sample Prep 100%
3.8 4.8 4.1 2.4 Overall = 3.8
5%
0%
Method
Avg. % Deviation from Mean**
Manual Automated
4%
2.1 1.7
2%
| xi,1 - xi | + | xi,2 - xi |
0% 1
5
9
13
17
21
25
29
33
37
Sample #
CONCLUSIONS
Automation System
■■
10%
Day 1 Day 3
Avg. % Variation*
**% of deviation from the mean =
2 xi
where xi =
xi,1 + xi,2 2
Figure 5: Comparison of sample deviation from mean in duplicate runs from both sample prep methods
Aliquot to filter plate
Elapsed Time (96 samples)
®
15%
Figure 3: Comparison of total THC measured by both sample prep methods
Place on robot deck
Add ISTD
6 hrs
20%
0%
Average deviation from mean
Original Samples
Aliquot supernatant to HPLC vial
25%
0%
% Total THC Automated
Concentrate Samples
Cannabinoid Analysis Workflows: Manual vs Automated
Flower
Day 2 Day 4
Day 1 Day 3
30%
20%
Figure 1: Comparison of total THC measured by both sample prep methods
METHODS
Manual
35%
Day
Variation of Measured Total THC in Plant: Manual vs. Automated Sample Prep
Actual Measured Total THC in Plant: Manual vs. Automated Sample Prep % Total THC Automated
Flower Samples
Automation of sample preparation procedures can help laboratories achieve higher throughput and also improve accuracy compared to manual methods. Recently, an automated liquid handling system was installed in the chemistry laboratory at RM3 Labs to aid with sample processing for potency analysis of cannabis plants and concentrates. Before incorporation of the instrument into routine analyses, the automated sample preparation method was compared to the standard manual method for validation. The validation study focused on total THC concentration in flowers (n = 88) and concentrates (n = 88) prepared by both the existing manual method and the automated method, and measurements were performed on four separate days to ensure reproducibility. A comparison of the data between the two methods showed good correlation with R2 values of > 0.94 for plants and > 0.97 for concentrates. Additionally, the mean relative percent difference between the two methods for accuracy and precision were both less than 5%. In addition to yielding satisfactory performance results, it is estimated that the automated method saves 4 – 5 hours/day of technician time.
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Automated sample preparation of cannabis flower (n = 88) and concentrate (n = 88) for cannabinoid analysis was shown to be comparable to manual methods. Based on internal acceptance criteria, the automated method was validated for routine use. According to a reproducibility study (n = 37), the automated method yielded a smaller average % deviation from the mean compared to the manual method. Automation can save an estimated 4 – 5 hours of preparation time for a batch of 96 samples.
ACKNOWLEDGMENTS CANNABIS SCIENCE CONFERENCE We would like to recognize all of the contributors at RM3 labs for performing the validation experiments and generating the data used for this poster.