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Can HPLC-UV Be Used For Terpenes Analysis In Cannabis?

By
  • Tim Herring
Tags
  • #Cannabinoids
  • #Terpenes
  • #Product Selection
  • #Raptor LC Columns
  • #Plant Tissue
  • #Method Development
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  • #Blogs
  • #Cannabis
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  • #Botanicals
  • #HPLC & UHPLC Columns
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While HPLC may be tempting to use for terpenes analysis, a GC/FID or GC/MS is really the most straightforward and recommended way of analyzing terpenes in cannabis. Terpenes, being relatively volatile and neutral, lend themselves rather nicely to GC in general.

As you can see from Figure 1 below, coelutions of the cannabinoids and terpenes are very likely when analyzing real cannabis samples by HPLC-UV methods. A Shimadzu Prominence 20AD HPLC system and Raptor ARC-18, 2.7um, 150 x 4.6mmID column (catalog# 9314A65) was used in Figure 1.

Figure 1- Standards at equal concentrations
figure-1-blog-terpenes

At 205nm, there are many co-elutions of terpenes and the cannabinoids, making identification and quantitation extremely difficult, if not impossible, for either class of compounds.

Conversely, cannabinoids can be analyzed by HPLC-UV, as long as the correct UV wavelength is chosen. At 220nm, terpenes yield a very low signal. Because cannabinoids of interest are present at a much higher concentration than terpenes, in addition to providing a stronger UV signal at 220nm, cannabinoids can be reliably analyzed at a wavelength of 220nm. See Figure 2.

Figure 2-

ColumnRaptor ARC-18 (cat.# 9314A65)
Dimensions:150 mm x 4.6 mm ID
Particle Size:2.7 µm
Pore Size:90 Å
Guard Column:Raptor ARC-18 EXP guard column cartridge 2.7 µm (cat.# 9314A0250)
Temp.:30 °C
SampleTetrahydrocannabivarin (cat.# 34100)
Cannabidiolic acid (CBDA) (cat.# 34094)
Cannabichromene (CBC) (cat.# 34092)
Cannabigerol (CBG) (cat.# 34091)
delta-9-Tetrahydrocannabinolic acid A (THCA-A) (cat.# 34093)
delta-8-Tetrahydrocannabinol (Δ8-THC) (cat.# 34090)
delta-9-Tetrahydrocannabinol (Δ9-THC) (cat.# 34067)
Cannabinol (CBN) (cat.# 34010)
Cannabidiol (CBD) (cat.# 34011)
Compounds not present in these mixes were obtained separately.
Diluent:25:75 Water:methanol
Conc.:50 µg/mL
Inj. Vol.:5 µL
Mobile Phase
A:Water, 5 mM ammonium formate, 0.1% formic acid
B:Acetonitrile, 0.1% formic acid
Time (min)Flow (mL/min)%A%B
0.001.52575
9.001.52575
DetectorUV/Vis @ 228 nm
InstrumentHPLC

 

In summary, HPLC-UV analysis of terpenes in cannabis is not recommended, and will likely cause more issues than it will provide solutions.

A good solution to the coelutions by HPLC-UV is to choose a GC headspace method. Interferences from the complex sample matrix, as well as the much less volatile cannabinoids can be eliminated then.

 

 Contact us or your Restek sales representative if you have any questions.

 

Comments

Fri, Mar 22, 2019

These are almost exclusively monoterpenes and some sesquiterpens. Are the HPLC limitations also applicable to diterpenes and triterpenes ?

Wed, Apr 17, 2019

This would need to be determined experimentally. If a C18 is too retentive, then perhaps try a C8 next, for instance.

Fri, Jan 06, 2017

Hello Amit, Unfortunately we have no data to say otherwise. I would surmise that ion suppression will result with the complex sample matrix. If compounds coelute, then that might be acceptable, as long as they are not isobaric. A biphenyl column could be tried, in addition to a standard C18. Selectivity can be greatly affected on the biphenyl, simply by changing mobile phase solvents (methanol vs acetontrile). Having said that, LC-MS (or LC-MS/MS) should be the best platform to do these simultaneously. I believe there is a company or two that has data on analyzing terpenes in cannabis products by LC-MS/MS. Thanks, Tim

Fri, Dec 30, 2016

Interesting work and good discussion Tim. Thank you! Do you think using LC-MS would provide enough selectivity to do Terpenes and Cannabinoid analysis together in one run??