No pressure! Low-Pressure Gas Chromatography Analysis of Cannabis Terpenes

Terpenes are responsible for the characteristic aromas and flavors of different natural and consumer products. They produce a wide range of scents, from citrusy and fruity to earthy and piney. Terpenes can be extracted from plant materials and used in various products, including essential oils, aromatherapy, and even as flavor additives in food and beverages. They are also an important part of cannabis' aroma and flavor.

The specific terpene profile of a cannabis strain is influenced by its genetics and growing conditions. Different strains can have vastly different terpene profiles. In the cannabis industry, terpene testing is often conducted alongside cannabinoid testing to provide consumers with information about a strain's terpene content.

In the analysis of cannabis terpenes, utilizing a conventional GC-MS approach is a commonplace technique. However, analysts can harness a powerful technique to reduce run times, increase sample throughput, and reduce lab costs: low-pressure gas chromatography (LPGC). LPGC is a technique that employs sub-ambient pressure conditions to create a vacuum system, while using a short, narrow restrictor column at the injection port, connected by a union to the actual analytical column. Generally, the analytical column will be 0.32 mm or wider. In this case, the analytical column of choice was the Rxi-624Sil MS column. This column is a great phase choice for this analysis because it is low-bleed, selective, and efficient.

The speed of analysis and increased sample volume are the major benefits of LPGC. Peak shapes are comparable to conventional GC-MS, with LPGC peak heights often being higher. The principal drawback of using LPGC is decreased separation efficiency. This can be problematic in the case of closely eluting isobars (compounds that cannot be resolved using MS spectra).

	Peaks	tR (min)	Conc. (µg/mL)
1.	α-Pinene	3.309	5
2.	Camphene	3.438	5
3.	β-Myrcene	3.627	5
4.	β-Pinene	3.639	5
5.	δ-3-Carene	3.810	5
6.	α-Terpinene	3.885	5
7.	α-Ocimene	3.969	5
8.	D-Limonene	3.969	5
9.	p-Cymene	3.996	5
10.	β-Ocimene	4.074	5
11.	Eucalyptol	4.074	5

	Peaks	tR (min)	Conc. (µg/mL)
12.	γ-Terpinene	4.197	5
13.	Terpinolene	4.468	5
14.	Linalool	4.882	5
15.	Isopulegol	5.632	5
16.	Geraniol	7.139	5
17.	β-Caryophyllene	7.989	5
18.	α-Humulene	8.127	5
19.	Nerolidol 1	8.373	5
20.	Nerolidol 2	8.460	5
21.	Guaiol	8.634	5
22.	Caryophyllene oxide	8.658	5
23.	α-Bisabolol	8.826	5

α-Ocimene and β-Ocimene have coeluted with D-Limonene and Eucalyptol, respectively.

Column	LPGC Rxi-624Sil MS (cat.# 11804), includes Rxi-624Sil MS, 10 m, 0.32 mm ID, 1.8 μm w/5 m, 0.15 mm ID Rxi restrictor connected via SilTite connector
Standard/Sample
	Cannabis terpenes standard #1 (cat.# 34095)
	Cannabis terpenes standard #2 (cat.# 34096)
Diluent:	Isopropanol
Conc.:	25 µg/mL (5 µg/mL on column)
Injection
Inj. Vol.:	1 µL split (split ratio 5:1)
Liner:	Topaz, splitless, single taper inlet liner, 4.0 mm x 6.5 x 78.5 (cat.# 23303)
Inj. Temp.:	280 °C
Oven
Oven Temp.:	55 °C (hold 1 min) to 130 °C at 30 °C/min (hold 3 min) to 280 °C at 60 °C/min (hold 2 min)
Carrier Gas	He, constant flow
Flow Rate:	0.9 mL/min

Detector	Agilent 5975C
Scan Program:
Group Start Time (min) Scan Range (amu) Scan Rate (scans/sec) 1 1.50 35 - 280
Transfer Line Temp.:	310 °C
Source Temp.:	330 °C
Quad Temp.:	180 °C
Solvent Delay Time:	1.50 min
Tune Type:	PFTBA
Ionization Mode:	EI
Instrument	Agilent 7890A GC & 5975C MSD
Sample Preparation	The sample was in a 2 mL short-cap, screw-thread vial (cat.# 21143) and capped with a short-cap, screw-vial closure (cat.# 24495).

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The conventional GC-MS run takes about 16 minutes with the latest compound eluting around 15 minutes. With LPGC, analysis time drops to 11 minutes. An analyst can note by using this method, there is an opportunity for great time and helium savings (as the method is shorter and operating under lower flow conditions.)

LPGC provided excellent peaks for those compounds that were fully resolved. Peaks were taller than those on the conventional chromatogram comparison. Of the compounds in the standard, beta-pinene, beta-myrcene, D-limonene, p-cymene, guaiol, and caryophyllene oxide were the compounds which suffered the most from the lack of separation efficiency. Beta-pinene and beta-myrcene coelution is the most concerning due to being isobars and very abundant in cannabis samples. Ocimene 1 and 2 were not detected in the initial run, however an analyst may identify their presence through mass spectrometry confirmation. Ocimene 1 and 2 seem to coelute with D-limonene and eucalyptol, respectively.

Because of the nature of terpenes in cannabis analysis, a higher chromatographic efficiency is paramount to high-quality results. As part of the same class of compounds, the spectra of terpenes can mimic each other very closely. An analyst could be faced with issues differentiating and quantitating compounds even after extracting chromatogram ions. LPGC is great for fast analysis. Analysts may find that LPGC is a great screening tool for cannabis samples.

Below is a comparison of conventional GC-MS analysis conditions of our cannabis terpenes standards with the LPGC analysis conditions.