No.

SSI-GCMS-1604

Gas Chromatograph Mass Spectrometer

Simplified Cannabis Terpene Profiling

by GCMS
No. GCMS-1604
Introduction
Terpene and terpenoid compounds are naturally
occurring aromatic compounds that give cannabis its
unique flavor and fragrance. Aside from their
aromatic properties, terpenes have advantageous
health benefits. They also have a synergistic
relationship with cannabinoids, which further
enhance the therapeutic effect of THC.
Cannabis has over 140 terpene components, many
of which are of medicinal interest.1 Predominant
terpenes in cannabis include -myrcene, which has
antibiotic properties and enhances the THC muscle
relaxant effect; -pinene, which has antiinflammatory properties and enhances the THC
bronchodilator effect; and -caryophyllene, which
also has anti-inflammatory properties and enhances
the THC gastric cytoprotective effect amongst other
health benefits. 2, 3 The concentration of individual
terpenes varies by strain, can be anywhere from 0.1
to 1.5% of its total dry weight, and can vary
depending on harvest time, drying and storage
conditions.1, 4 Terpene levels can decrease over time,
and after three months of storage, can reduce
terpene levels by more than half.4 The decrease in
terpene amount over time varies for different
terpenes.

accordance with NRS 453A.368 in May of 2015.5

Due to the uniqueness of terpene profiles, they can
be used by cultivators as a fingerprint to partially
ID the specific strain in question. This application
note describes the analysis of several strains of
cannabis for 41 terpenes on Gas Chromatography
Mass Spectrometry (GCMS) with headspace
injection.
Experimental
Terpene analyses were conducted using a Shimadzu
GCMS-QP2010SE single quadrupole mass
spectrometer with the HS-20 headspace autosampler
for sample introduction. As terpene concentration
can be in the percent level, effluent from the HS-20
was split 50-to-1 to allow for analyzing high
concentration standards. The MS was run using the
FAAST (Fast Automated SCAN/SIM Type) where in
the SCAN mode was used for identification and the
SIM mode was used for greater quantitation. The
instrument and operating conditions are shown in
Table 1.

Recent proliferation of new terpene profiling

methods can be attributed to the ever-increasing
state legalization of cannabis use. The state of
Nevada added the testing of ten terpenes in
Shimadzu GCMS-QP2010SE

No. SSI-GCMS-1604

Table 1: Instrument Operating Conditions and Method Parameters

Head Space
Operation Mode
Sample
Equilibration
Sample Loop
Sample Pathway Temperature
Transfer Line Temperature
Gas Chromatogram
Injection
Column

Oven Program

Detector
Operating mode
Ion Source
Solvent Cut Time
MS Interface

HS-20 Loop Model

Static headspace with loop
10uL sample volume
10-mL headspace vial
30 minutes at 150oC
1-mL loop
Vial pressurization 1.00 min, equilibration 0.20 min
Loop load time 1.00 min, equilibration 0.20 min
Injection time 1.0 min
150oC
150oC
GC-2010 Plus
Split injection from HS-20, with 50:1 split ratio
Rxi-624 Sil MS 30.0m x 0.25 mm x 1.40 m
Helium carrier gas
Constant linear velocity, 47.2 cm/sec
Column Flow 1.64 mL/min
Purge Flow 3.0 ml/min
80 oC, hold 1.0 min
12 oC /min to 150 oC, hold 1.0 min
9 oC /min to 250 oC, hold 1 min
Total GC run time 19.94 min
Total cycle time 24 min
GCMS-QP2010 SE
Selected Ion Monitoring (SIM) and SCAN
200 oC, EI mode, 70eV
2 min
300 oC

Sample Preparation
In order to obtain a more complete terpene profile,
two sets of standards were used. Standard one,
purchased from SPEX (Metuchen, NJ), included 38
terpenes in a 100g/mL stock solution. Standard
two, purchased from Restek (Bellefonte, PA),
included 23 terpenes in a 2500 g/mL stock solution.
As plant material does not dissolve in solvent, the full
evaporation headspace technique (FET) is used for
quantitation. Using FET, a small amount of standard
and sample is used to create a single phase gas
system, as compared to a two phase liquid-gas
system as in traditional headspace techniques.6 A
five-point calibration curve was created from the
SPEX terpene standard with concentrations ranging
from 12.5-100 g/mL. An aliquot of 10L of the
standard was placed in a 10mL headspace vial and
capped. A seven-point calibration curve was created
from the Restek standard in a similar fashion as
described above, with the concentration range from
78.125 g/mL to 2500 g/mL. The calculated
amount of terpene injected on column is shown in
Table 2.

Table 2: Calculated amount of terpene standard on column

Standard
Concentration Injection Amount On(g/mL)
Volume
Column (g)
(mL)
SPEX
12.5
0.01
0.125
SPEX
25
0.01
0.25
SPEX
50
0.01
0.50
SPEX
75
0.01
0.75
SPEX
100
0.01
1.0
Restek
78.125
0.01
0.78
Restek
156.25
0.01
1.25
Restek
312.5
0.01
3.25
Restek
625
0.01
6.25
Restek
1250
0.01
12.5
Restek
2000
0.01
20.0
Restek
2500
0.01
25.0

A part of the flower weighing 1.0 gram was frozen,

followed by grinding to ensure a representative
sample. Ten to 30mg of the flower were then
weighed into a headspace vial and capped. The final
result was calculated to give wt %.

No. SSI-GCMS-1604

Results and Discussion

Chromatography
Figures 1a and 1b show the TIC chromatographic
separation using the SPEX and Restek standards.
(x100,000)
7.0
6.0
5.0
4.0
3.0
2.0
1.0
7.0

6.0

9.0

8.0

10.0

11.0

12.0

13.0

14.0

15.0

16.0

17.0

18.0

10.0

11.0

12.0

13.0

14.0

15.0

16.0

17.0

18.0

Figure 1a: TIC from SPEX standard

(x1,000,000)
2.5

2.0

1.5

1.0

0.5

7.0

6.0

8.0

9.0

Figure 1b: TIC chromatogram from Restek standard

Calibration
Calibration standards were prepared and analyzed
using the optimized HS-20 and GC-MS parameters
with concentrations ranging from 0.125g to 1.0g
using the SPEX terpene standard, and 0.78g to
25g using the Restek standard. All points on the
calibration curve were run in replicates of six. Figure
2 shows the terpene calibration curves required by

the state of Nevada with corresponding correlation

coefficients illustrating linearity. The SIM
chromatograms represent the lowest calibration
point (0.78g) for each terpene. All of the
chromatographic peaks were easily integrated at
each level. Full statistics on all calibration points are
listed in Table 3a and 3b.

Alpha-Pinene
Area(x1,000,000)

6.051

(x100,000)
93.00
1.5 91.00
92.00

7.5

1.0

5.0
0.5

2.5

6.1

6.0

5.9

0.0
0.0

6.2

5.0

10.0

15.0

20.0

Conc.

20.0

Conc.

R2 = 0.999
Beta-Mycene
Area(x1,000,000)
7.5

6.726

(x100,000)
93.00
1.5 41.00
69.00
1.0

5.0

0.5

2.5

6.6

6.7

6.8

6.9

0.0
0.0

5.0

10.0

15.0

R2 = 0.999

No. SSI-GCMS-1604

Beta-Pinene
Area(x1,000,000)

6.779

(x100,000)
93.00
1.5 69.00

7.5

1.0

5.0
0.5

2.5

6.7

6.6

0.0
0.0

6.9

6.8

5.0

10.0

15.0

20.0

Conc.

20.0

Conc.

20.0

Conc.

20.0

Conc.

R2 = 0.999
alpha.-Terpinene
Area(x1,000,000)

7.290

(x100,000)
121.00
1.25 93.00
136.00
1.00

5.0

0.75

2.5

0.50
0.25

0.0
0.0

7.5

7.4

7.3

7.2

5.0

10.0

15.0

R2 = 0.999
(R)-(+)-Limonene
Area(x1,000,000)

7.460

(x100,000)
68.00
1.00 67.00
93.00

5.0

0.75
0.50

2.5
0.25

7.4

7.3

0.0
0.0

7.6

7.5

5.0

10.0

15.0

R2 = 0.999
Linolool
Area(x1,000,000)

9.002

(x10,000)
71.00
93.00
55.00
5.0

2.5
2.5

8.9

9.0

9.1

9.2

0.0
0.0

5.0

10.0

15.0

R2 = 0.999

No. SSI-GCMS-1604

Beta-Caryophyllene
Area(x1,000,000)
3.0

14.422

(x10,000)
93.00
4.0 91.00
3.0

2.0
2.0

1.0

1.0

14.6

14.5

14.4

14.3

0.0
0.0

5.0

10.0

15.0

20.0

Conc.

20.0

Conc.

20.0

Conc.

20.0

Conc.

R2 = 0.999
alpha.-Humulene
Area(x1,000,000)

14.972

(x100,000)
93.00
80.00
1.00 121.00

7.5

0.75

5.0
0.50

2.5

0.25

0.0
0.0

15.1

15.0

14.9

14.8

5.0

10.0

15.0

R2 = 0.999
Caryophyllene oxide
Area(x1,000,000)
17.372

(x10,000)
79.00
41.00
43.00

2.0

2.0

1.0
1.0

0.0
0.0
17.4

17.3

17.2

5.0

10.0

15.0

R2 = 0.997

17.5

Alpha-Bisabolol
Area(x1,000,000)

18.218

(x10,000)
43.00
119.00
5.0 69.00

3.0
2.0

2.5

1.0

18.1

18.2

18.3

18.4

0.0
0.0

5.0

10.0

15.0

R2 = 0.994
Figure 2: Calibration curves for Restek terpene testing required by the state of Nevada; ranges shown are 0.78g to 25g on column

No. SSI-GCMS-1604

Table 3a: 5-point calibration and precision results for analysis of terpenes (SPEX standards)
Calibration Results
RF RSD (%)
5.549
3.642
3.440
3.732
3.998
4.367
5.247
5.916
3.757
5.817

R2
0.998
0.999
0.999
0.997
0.998
0.999
0.999
0.996
0.998
0.997

Precision at 12.5ppm
(n=6)
Mean Conc
RSD (%)
0.123
0.796
0.124
1.164
0.121
1.447
0.126
1.317
0.124
1.180
0.124
3.533
0.124
3.386
0.107
1.269
0.125
1.041
0.123
0.887

4.109

0.997

0.124

1.334

0.950

2.776

3.675
3.695
4.095
9.252
2.863
0.607

0.998
0.998
0.998
0.995
0.999
0.999

0.124
0.124
0.124
0.122
0.126
0.125

0.766
0.908
1.024
2.926
1.763
1.011

0.952
0.947
0.948
0.950
0.989
-

2.720
1.954
1.426
5.300
2.055
-

5.338

0.999

0.125

4.560

0.980

0.979

2.433
0.785
8.659
4.979
4.774
1.376
2.161
11.23
14.27
1.859
3.574
3.058
2.899
2.146
2.382
3.688
1.577
1.565
3.077
2.911

0.999
0.999
0.992
0.998
0.997
0.999
0.999
0.998
0.999
0.999
0.998
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.999

0.126
0.125
0.130
0.128
0.125
0.125
0.126
0.127
0.125
0.124
0.126
0.124
0.126
0.125
0.126
0.126
0.125
0.125
0.126
0.126

2.210
0.952
6.949
3.287
5.422
1.909
1.657
6.418
8.386
2.093
0.922
1.087
1.899
1.180
1.427
1.141
5.449
1.246
1.211
1.862

0.981
1.050
1.005
0.983
0.981
0.985
0.997
0.985
0.973
0.979
0.958
1.021
0.969
0.979
0.993
0.988
0.976
0.984
0.986

2.109
2.745
1.939
2.094
1.242
1.359
3.828
2.858
1.841
2.142
1.630
4.524
1.652
2.107
2.531
1.360
1.237
1.750
1.536

Terpene
Alpha-Pinene
Camphene
Sabinene
beta.-Myrcene
beta.-Pinene
.alpha.-Phellandrene
(1S)-3-Carene
alpha.-Terpinene
trans-.beta.-Ocimene
(R)-(+)-Limonene
Ocimene (mixture of
isomers)
Eucalyptol
gamma.-Terpinene
Terpinolene
Sabedine Hydrate
Linolool
Fenchone (mix of
isomers)*
(R)-Endo-(+)-Fenchyl
Alcohol
Isopulegol
Camphor (mix of isomers)*
Isoborneol
Hexahydrothymol
Borneol (isomers)
Terpineol (mix of isomers)
gamma.-Terpineol
Nerol
Geraniol
Pulegone
Geraniol acetate
alpha.-Cedrene
Trans-Caryophyllene
alpha.-Humulene
Valencene
cis-Nerolidol
trans-Nerolidol
Guaiol
(+)-Cedrol
.alpha.-Bisabolol

* Data at the high level is not applicable

Precision at 100 ppm

(n=6)
Mean Conc
RSD (%)
0.948
2.980
0.964
3.734
0.972
2.005
0.994
3.038
0.963
2.294
0.959
2.602
0.978
2.841
0.935
1.982
0.957
2.857
0.950
1.914

No. SSI-GCMS-1604

Table 3b: 7-point calibration and precision results for analysis of terpenes (SPEX standards)
Calibration Results
RF RSD (%)
2.545
2.095
1.833
1.973
2.029
2.150
1.746
1.611
2.777

R2
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.999

Precision at 78.125 ppm

(n=6)
Mean Conc
RSD (%)
0.764
1.478
0.766
1.674
0.768
1.599
0.767
1.386
0.767
1.412
0.766
1.611
0.777
1.349
0.777
1.630
0.767
1.780

1.452

0.999

0.772

1.395

25.011

2.494

3.597
2.231
1.831
3.545
2.816
10.282
2.077
2.213
2.532
3.043
5.176
9.397
9.873

0.998
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.999
0.996
0.997
0.993

0.768
0.766
0.768
0.784
0.780
0.796
0.768
0.768
0.772
0.769
0.757
0.804
0.046

1.498
1.562
1.553
1.156
1.773
1.273
1.493
1.430
1.640
1.699
1.535
7.127
28.412

25.664
24.578
24.726
25.783
25.432
25.944
24.565
24.436
24.095
23.772
22.852
26.803
24.053

4.007
2.489
2.380
1.876
1.992
2.812
2.487
2.553
3.597
3.692
3.800
4.869
3.741

Terpene
Alpha-Pinene
Camphene
beta.-Myrcene
beta.-Pinene
(1S)-3-Carene
alpha.-Terpinene
trans-.beta.-Ocimene
(R)-(+)-Limonene
p-cymene
Ocimene (mixture of
isomers)
Eucalyptol
gamma.-Terpinene
Terpinolene
Linolool
Isopulegol
Geraniol
Trans-Caryophyllene
alpha.-Humulene
cis-Nerolidol
trans-Nerolidol
Guaiol
Caryophyllene oxide
.alpha.-Bisabolol

Cannabis Results
Table 4 shows the wt% of terpenes in three different
strains of cannabis: CB Diesel, Blue Dream, and Haze
Wreck.

Precision at 2500 ppm

(n=6)
Mean Conc
RSD (%)
24.614
2.12
24.501
2.195
24.604
3.013
24.725
1.948
24.690
2.390
24.706
1.940
25.381
2.525
25.395
1.914
24.712
2.390

Figure 5 shows the chromatograms of the required

terpenes by the state of Nevada.

Table 4: wt% of terpenes in three different strains of cannabis

0.5
0.45
0.4
0.35
0.3
0.25
0.2
0.15
0.1
0.05
0

alpha.-Pinene
Camphene
Sabinene
.beta.-Myrcene
beta.-Pinene
.alpha.-Phellandrene
(1S)-3-Carene
alpha.-Terpinene
trans-.beta.-Ocimene
(R)-(+)-Limonene
p-cymene
Ocimene (mixture of
Eucalyptol
gamma.-Terpinene
Terpinolene
Sabedine Hydrate
Linolool
Fenchone (mis of
(R)-Endo-(+)-Fenchyl
Isopulegol
Camphor (mix of
Isoborneol
Hexahydrothymol
Borneol (isomers)
Terpineol (mix of
gamma.-Terpineol
Nerol
Geraniol
Pulegone
Geraniol acetate
alpha.-Cedrene
Trans-Caryophyllene
Beta-Caryophyllene
alpha.-Humulene
Valencene
cis-Nerolidol
trans-Nerolidol
Guaiol
Caryophyllene oxide
(+)-Cedrol
.alpha.-Bisabolol

Wt% of Terpenes in Cannabis

CB Diesel

Blue Dream

Haze Wreck

No. SSI-GCMS-1604

Figure 5: Chromatograms for terpenes required by the state of Nevada for three strains of cannabis

Blue Dream

Haze Wreck
(x1,000,000)
3.0 93.00
91.00
92.00
2.0

6.046

(x1,000,000)
1.00 93.00
91.00
0.75 92.00

6.052

CB Diesel
(x1,000,000)
3.0 93.00
91.00
92.00
2.0

6.052

Terpene
Alpha-Pinene

0.50

6.2

6.1

6.0

6.1

(x1,000,000)
1.5 93.00
41.00
69.00
1.0

6.727

(x1,000,000)
93.00
5.0 41.00
69.00

6.2

6.1

6.0
(x1,000,000)
2.0 93.00
41.00
69.00

6.721

6.0

Beta-Mycene

1.0

0.25

6.2

6.730

1.0

1.0

2.5
0.5

6.6

Beta-Pinene

6.7

6.8

6.6

6.7

6.8

6.6

(x1,000,000)
1.5 93.00
69.00

(x1,000,000)
93.00
5.0 69.00

6.7

6.8

(x1,000,000)
2.0 93.00
69.00

6.7

(x10,000)
7.5 121.00
93.00
136.00
5.0

(x10,000)
121.00
93.00
1.0 136.00

2.5

0.5

7.3

1.0

6.782

6.9

6.7

6.8

6.9

(x10,000)
121.00
7.5 93.00
136.00
5.0
2.5

7.4

7.2

7.3

(x1,000,000)
1.0 68.00
67.00
93.00

7.461

(x1,000,000)
68.00
1.5 67.00
93.00

6.8

7.2

7.4

7.3

(x1,000,000)
68.00
67.00
1.0 93.00

0.5

7.4

7.465

7.2

(R)-(+)Limonene

6.9

1.0

7.286

6.8

7.291

alpha.Terpinene

0.5

7.455

6.7

6.774

6.779

1.0
2.5

0.5

0.5
0.0
7.4
(x100,000)
71.00
5.0 93.00
55.00

9.004

(x100,000)
71.00
93.00
55.00

7.6

7.5

7.6

7.4

7.5

(x100,000)
71.00
93.00
2.0 55.00

7.6

9.007

Linolool

7.5

8.998

7.4

2.5
2.5

0.0

BetaCaryophyllene

9.0

5.0

8.9

9.0

(x100,000)
5.0 93.00
91.00

14.427

(x100,000)
93.00
91.00

9.1

9.1

8.9

9.0

(x100,000)
93.00
91.00

14.418

8.9

9.1

14.429

0.0

1.0

2.5

2.5

2.5

14.3

14.4

14.5

14.3

14.4

14.5

0.0
14.3

14.4

14.5

No. SSI-GCMS-1604

(x100,000)
93.00
80.00
3.0 121.00

2.5

2.5

14.978

(x100,000)
93.00
80.00
121.00

14.968

(x100,000)
5.0 93.00
80.00
121.00

14.974

alpha.Humulene

2.0
1.0

0.0
15.0

14.9

Caryophyllene
oxide

15.1

14.9

(x1,000)
79.00
41.00
5.0 43.00

15.1

15.0

5.0
2.5

14.9

15.0

15.1

17.4

17.5

(x1,000)
7.5 79.00
41.00
43.00
5.0

(x1,000)
79.00
7.5 41.00
43.00

2.5

2.5
0.0

AlphaBisabolol

17.5

17.4

17.4

(x10,000)
3.0 43.00
119.00
69.00
2.0

18.223

(x10,000)
43.00
119.00
69.00

17.3

2.5

17.5

1.0
0.1
18.1

18.2

0.0
18.1

18.3

Conclusion
The first sample of cannabis analyzed, CB Diesel, was
analyzed shortly after harvest. The resulting wt% of
terpenes is similar to that in current literature.1 The
other two samples, Blue Dream and Haze Wreck,
were stored at ambient temperature and exposed to
light for one month prior to analyzing. It has been
demonstrated that different storage conditions can

17.3
(x10,000)
1.5 43.00
119.00
69.00
1.0

18.217

17.3

0.5

18.2

18.3

18.1

18.2

18.3

change terpene results over time,4 and this should be

taken into consideration when analyzing cannabis
samples as the results show less than expected
results. Varying storage conditions and degradation
experiments should be the next study in the everchanging world of regulatory cannabis testing.

References
1. Giese, M.W., Lewis, M.A., Giese, L., and Smith, K.M. Development and Validation of a Reliable and
Robust Method for the Analysis of Cannabinoids and Terpenes in Cannabis. Napro Research, California,
2015.
2. Parland, J.M., and Russo, E.B. Cannabis and Cannabis Extracts: Greater than the Sum of Their Parts? The
Haworth Press, Pennsylvania, 2001
3. [1] E. Russo, Taming THC: Potential Cannabis Synergy and Phytocannabinoid-Terpenoid Entourage Effects,
British Journal of Pharmacology 163 (2011) 1344.
4. Chemistry and Analysis of Phytocannabinoids and Other Cannabis Constituents. Humana Press, New
Jersey
5. Information on all Policies related to Nevadas Medical Marijuana Establishments can be found at
http://dpbh.nv.gov/Reg/MME/MME_-_Home/
6. http://www.restek.com/Technical-Resources/Technical-Library/Foods-Flavors-Fragrances/fff_FFAN2045-UNV
For more information on Shimadzus solutions for cannabis testing, please visit www.GrowYourLab.com

First Edition: September 2016

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