Essential Oils using GC-VUV An Automated, Speed-based Approach Alex - - PowerPoint PPT Presentation

▶
essential oils using gc vuv
SMART_READER_LITE
LIVE PREVIEW

Essential Oils using GC-VUV An Automated, Speed-based Approach Alex - - PowerPoint PPT Presentation

Jan 01, 2023 146 likes •500 views

Terpenes Analysis of Essential Oils using GC-VUV An Automated, Speed-based Approach Alex Hodgson, Senior Applications Chemist VUV Analytics, Inc. VUV Absorption Spectroscopy Simple absorption spectroscopy in a new spectral region Vacuum

slide-1
SLIDE 1

Terpenes Analysis of Essential Oils using GC-VUV

An Automated, Speed-based Approach

Alex Hodgson, Senior Applications Chemist VUV Analytics, Inc.

slide-2
SLIDE 2

VUV Absorption Spectroscopy

  • Simple absorption spectroscopy in a new spectral region
  • Vacuum ultraviolet (VUV) spectroscopy is a universal technique that

provides unique spectral fingerprints

  • High energy, low wavelength exposure produces electronic transitions

between σ→σ*, n→σ*, and π→π* molecular orbitals

  • Compound identification is unambiguous, even for structural isomers
  • Chromatographic compression leads to higher sample throughput
  • Software deconvolution of coelutions reduces priority on chromatographic

separation

  • Chromatography runs can be deliberately compressed
slide-3
SLIDE 3
  • Class of organic compounds produced by plants (and some insects)
  • Primary constituents of essential oils, which contribute heavily to

fragrances and flavors

  • Widely

utilized in the food and fragrance, cosmetics, and pharmaceuticals industries

  • Composed of multiple units of isoprene (C5H8)
  • Monoterpenes – 2 isoprene units (C10H16)
  • Sesquiterpenes – 3 isoprene units (C15H24)
  • Very high prevalence of structural isomers

Introduction to Terpenes

slide-4
SLIDE 4

Monoterpenes Monoterpenoids

α-Pinene β-Myrcene Eucalyptol Linalool β-Caryophyllene α-Humulene trans-Nerolidol

Sesquiterpenes Sesquiterpenoids

p-Cymene Limonene Guaiol

slide-5
SLIDE 5

Headspace/Solid-phase Microextraction (SPME) Setup

Agilent 6890 Gas Chromatograph VUV Analytics VGA-100 Gerstel MPS2 Autosampler

slide-6
SLIDE 6
  • Gerstel MPS2
  • Incubator: 80°C for 10 minutes, 250 rpm agitation (10 sec on, 1 sec off)
  • Syringe: 90°C, 2.5 mL injection volume
  • Agilent 6890 GC
  • Inlet: 250°C, split 2.5:1
  • Column: Restek Rxi-624Sil MS (30 m x 0.25 mm, 1.40-μm)
  • Flow: 4 mL/min helium, constant flow
  • Oven: 60°C, hold 0.1 min; 23.8°C/min to 300°C (run time - 10.2 min)
  • VUV Analytics VGA-100
  • Makeup gas: 0.25 psi N2
  • Flow cell and transfer line: 275°C
  • Acquisition rate: 4 spectra/sec
  • Acquisition range: 125-240 nm

Headspace Run Conditions

slide-7
SLIDE 7

Terpenes Standard (Restek)

Class Compound RT (min) Formula MW Sources Monoterpene α-Pinene 3.87 C10H16 136.23 Coniferous trees, rosemary, eucalyptus Camphene 4.03 C10H16 136.23 Camphor, neroli, valerian β-Myrcene 4.21 C10H16 136.23 Thyme, cardamom, hops, cannabis β-Pinene 4.25 C10H16 136.23 Nutmeg, rosemary, sage 3-Carene 4.43 C10H16 136.23 Turpentine, rosemary, cedar α-Terpinene 4.50 C10H16 136.23 Allspice, juniper, marjoram cis-Ocimene 4.53 C10H16 136.23 Basil, lavender, clary sage Limonene 4.58 C10H16 136.23 Citrus fruits, mint trans-Ocimene 4.64 C10H16 136.23 Basil, lavender, clary sage γ-Terpinene 4.77 C10H16 136.23 Citrus fruits, cumin, Syrian oregano Terpinolene 4.98 C10H16 136.23 Allspice, citrus fruits, juniper

slide-8
SLIDE 8

Terpenes Standard (Restek)

Class Compound RT (min) Formula MW Sources Monoterpenoid p-Cymene 4.60 C10H14 134.22 Cumin, thyme Linalool 5.21 C10H18O 154.25 Mint, cinnamon, rosewood Isopulegol 5.67 C10H18O 154.25 European pennyroyal, corn mint Geraniol 6.27 C10H18O 154.25 Almonds, palmarosa, rose Sesquiterpene β-Caryophyllene 7.32 C15H24 204.35 Cloves, rosemary, hops, cannabis α-Humulene 7.53 C15H24 204.35 Sage, ginseng, tobacco Sesquiterpenoid cis-Nerolidol 7.85 C15H26O 222.37 Neroli, jasmine, lavender trans-Nerolidol 8.01 C15H26O 222.37 Neroli, jasmine, lavender Guaiol 8.37 C15H26O 222.37 Cypress, pine, cannabis α-Bisabolol 8.73 C15H26O 222.37 German chamomile, balsam poplar

slide-9
SLIDE 9

0.1 0.2 0.3 0.4 0.5 3.5 4 4.5 5 5.5 6 6.5 7 7.5 8 8.5 9 Detector Response Time (min)

  • 1. α-Pinene
  • 12. Terpinolene
  • 2. Camphene
  • 13. Linalool
  • 3. β-Myrcene
  • 14. Isopulegol
  • 4. β-Pinene
  • 15. Geraniol
  • 5. 3-Carene
  • 16. β-Caryophyllene
  • 6. α-Terpinene
  • 17. α-Humulene
  • 7. cis-Ocimene
  • 18. cis-Nerolidol
  • 8. Limonene
  • 19. trans-Nerolidol
  • 9. p-Cymene
  • 20. Guaiol
  • 10. trans-Ocimene
  • 21. α-Bisabolol
  • 11. γ-Terpinene

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

125-160 nm 170-240 nm Last analyte elutes before 9 min!

slide-10
SLIDE 10

Mass Spectra of Coeluting Isomers

β-Myrcene β-Pinene

41 41 69 69 93 93 121 121 136 136

slide-11
SLIDE 11

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 130 140 150 160 170 180 190 200 210 220 230 240 Normalized Absorbance Wavelength (nm)

Absorbance Spectra of Monoterpene Isomers

α-Pinene β-Pinene β-Myrcene Limonene cis-Ocimene trans-Ocimene

slide-12
SLIDE 12

Spectral Deconvolution of Coeluting Peaks

Summary:

  • XXX
  • XXX
  • XXX

0.05 0.1 0.15 0.2 0.25 0.3 0.35 0.4 4.44 4.46 4.48 4.5 4.52 4.54 4.56 4.58 4.6 4.62 4.64 4.66 Detector Response Time (min)

125-240 nm Limonene α-Terpinene p-Cymene cis-Ocimene trans-Ocimene

slide-13
SLIDE 13

Region 1 – Single Analyte Spectral Matching

0.02 0.04 0.06 0.08 0.1 0.12 0.14 0.16 0.18 0.2 0.22 130 140 150 160 170 180 190 200 210 220 230 240 Absorbance Wavelength (nm)

Summed Retention Region Spectrum Summed Target Spectrum Best Fit; R^2 = 0.99890 alpha-Terpinene

slide-14
SLIDE 14

Region 2 – Summation of Spectra from Coelution #1

0.03 0.06 0.09 0.12 0.15 0.18 0.21 0.24 0.27 0.3 0.33 130 140 150 160 170 180 190 200 210 220 230 240 Absorbance Wavelength (nm)

Summed Retention Region Spectrum Summed Target Spectra Best Fit; R^2 = 0.99923 alpha-Terpinene cis-Ocimene

slide-15
SLIDE 15

Region 3 – Summation of Spectra from Coelution #2

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1.1 1.2 130 140 150 160 170 180 190 200 210 220 230 240 Absorbance Wavelength (nm)

Summed Retention Region Spectrum Summed Target Spectra Best Fit; R^2 = 0.99962 Limonene p-Cymene

slide-16
SLIDE 16

Region 4 – Single Analyte Spectral Matching

0.05 0.1 0.15 0.2 0.25 0.3 0.35 0.4 0.45 0.5 130 140 150 160 170 180 190 200 210 220 230 240 Absorbance Wavelength (nm)

Summed Retention Region Spectrum Summed Target Spectrum Best Fit; R^2 = 0.99771 trans-Ocimene

slide-17
SLIDE 17

VUV vs MS – Tale of the Tape

Who has the edge? Higher flow limit Lower relative LOD Ease of use/ Ease of maintenance 3-dimensional information Unique spectra for all measurable compounds Isomer differentiation

slide-18
SLIDE 18

VUV Methods – Run Conditions

Agilent 6890 GC

  • 1 µL injection
  • Inlet: 250°C, split 10-300:1
  • Column: Restek Rxi-1ms

(30 m x 0.25 mm, 0.25-µm)

VUV Analytics VGA-100

  • Makeup gas: 0.25 psi N2
  • Flow cell and transfer line: 275°C
  • Acquisition rate: 5 spectra/sec
  • Acquisition range: 125-240 nm

VUV Method 1 VUV Method 2 VUV Method 3 Col Flow (mL/min He) 1.4 2 4 Oven Start (°C) 40 40 40 Initial Hold (min) 0.2 0.2 0.1 Oven Ramp (°C/min) 9.6 20.4 30.9 Oven Final (°C) 250 250 250 Run Time (min)

22.1 10.5 6.85

slide-19
SLIDE 19

VUV Run Results

Peak # Analyte 1 α-Pinene 2 Camphene 3 β-Pinene 4 β-Myrcene 5 3-Carene 6 α-Terpinene 7 p-Cymene 8 Eucalyptol 9 Limonene 10 cis-Ocimene 11 trans-Ocimene 12 γ-Terpinene 13 Terpinolene 14 Linalool 15 Isopulegol 16 Geraniol 17 β-Caryophyllene 18 α-Humulene 19 cis-Nerolidol 20 trans-Nerolidol 21 Caryophyllene oxide 22 Guaiol 23 α-Bisabolol

slide-20
SLIDE 20

VUV Methods Comparison – Tea Tree Oil

Compound VUV Method 1 (mg/mL) – 22.1min VUV Method 2 (mg/mL) – 10.5min VUV Method 3 (mg/mL) – 6.85min VUV Average (mg/mL) CV (%)

α-Pinene 20.6 21.5 23.0 21.7 4.48 α-Terpinene 77.9 80.4 84.8 81.0 3.57 β-Caryophyllene 2.09 2.26 2.03 2.13 4.46 β-Myrcene 7.20 7.72 7.95 7.62 5.14 β-Pinene 5.83 5.85 5.21 5.63 5.30 Eucalyptol 21.0 25.0 22.8 22.9 7.24 γ-Terpinene 161 172 193 175 7.53 Limonene 6.38 5.62 7.03 6.34 9.08 p-Cymene 25.2 27.9 33.3 28.8 11.6 Terpinolene 27 29.7 23.1 26.6 10.3

slide-21
SLIDE 21

MS Methods – Run Conditions

Agilent 6890 GC

  • 1 µL injection
  • Inlet: 250°C, split 20-975:1
  • Column: Restek Rxi-1ms

(30 m x 0.25 mm, 0.25-µm)

  • Transfer line: 280°C

Agilent 5973N MS

  • EI Source: 300°C
  • Quad 1: 200°C
  • Scan range: 50-250

MS Method 1 MS Method 2 Col Flow (mL/min He) 1 1.4 Oven Start (°C) 40 50 Initial Hold (min) 1.5 1 Ramp 1 (°C/min) 1 15 Ramp 1 End (°C) 77

  • Ramp 2 (°C/min)

7

  • Oven Final (°C)

250 250 Run Time (min)

63.2 14.3

slide-22
SLIDE 22

MS Run Results

Peak # Analyte 1 α-Pinene 2 Camphene 3 β-Pinene 4 β-Myrcene 5 3-Carene 6 α-Terpinene 7 p-Cymene 8 Eucalyptol 9 Limonene 10 cis-Ocimene 11 trans-Ocimene 12 γ-Terpinene 13 Terpinolene 14 Linalool 15 Isopulegol 16 Geraniol 17 β-Caryophyllene 18 α-Humulene 19 cis-Nerolidol 20 trans-Nerolidol 21 Caryophyllene oxide 22 Guaiol 23 α-Bisabolol

slide-23
SLIDE 23

VUV vs MS Comparison – Tea Tree Oil

Compound MS Method 1 (mg/mL) – 63.2min Average w/ all VUV runs (mg/mL) CV (%) α-Pinene 26.3 22.8 9.56 α-Terpinene 91.2 83.6 6.04 β-Caryophyllene 1.58 1.99 12.6 β-Myrcene 7.44 7.58 3.76 β-Pinene 6.36 5.81 7.00 Eucalyptol 30.2 24.8 14.0 γ-Terpinene 205 183 9.48 Limonene 7.84 6.72 12.2 p-Cymene 35 30.4 13.1 Terpinolene 31.1 27.7 11.0

slide-24
SLIDE 24

MS Methods Comparison – Tea Tree Oil

Compound MS Method 1 (mg/mL) – 63.2min MS Method 2 (mg/mL) – 14.3min MS Average (mg/mL) CV (%) α-Pinene 26.3 27.3 26.8 1.75 α-Terpinene 91.2 96.2 93.7 2.71 β-Caryophyllene 1.58 2.39 1.99 20.4 β-Myrcene 7.44 9.78 8.61 13.6 β-Pinene 6.36 7.91 7.13 10.9 Eucalyptol 30.2 28.7 29.5 2.51 γ-Terpinene 205 202 204 0.788 Limonene 7.84 17.4 12.6 37.9 p-Cymene 35.0 43.0 39.0 10.2 Terpinolene 31.1 38.3 34.7 10.3

slide-25
SLIDE 25

Spectral Comparison – Limonene & Eucalyptol

1 125 150 175 200 225

Eucalyptol Limonene

m/z Norm Abund λ (nm) Norm Abs

slide-26
SLIDE 26

Class-Type Analysis of….Gasoline?

slide-27
SLIDE 27

Time Interval Deconvolution (TID)

  • Alternative quantitation method using VUV Analyze™ software
  • Chromatogram is divided into equal, small time intervals

(typically <0.05 min)

  • For each time interval, compare measured spectrum against

reference spectra in designated library, best analyte(s) fit determined

  • Can quickly be performed to measure total response per analyte

for a chromatogram; this can be converted into a relative mass percentage, relative volume percentage,

  • r

absolute concentration

slide-28
SLIDE 28

Essential Oil TID Run Conditions

  • Agilent 6890 GC
  • 1 μL injection
  • Inlet: 250°C, split 250:1
  • Column: Restek Rxi-1ms (30 m x 0.25 mm, 0.25-μm)
  • Flow: 2 mL/min helium, constant flow
  • Oven: 70°C, hold 0.1 min; 8.5°C/min to 250°C (run time – 21.2 min)
  • VUV Analytics VGA-100
  • Makeup gas: 0.25 psi N2
  • Flow cell and transfer line: 275°C
  • Acquisition rate: 10 spectra/sec
  • Acquisition range: 125-240 nm
slide-29
SLIDE 29

TID Analysis Using VUV Analyze™

α-Pinene Camphene β-Pinene β-Myrcene 3-Carene α-Terpinene p-Cymene Limonene cis-Ocimene trans-Ocimene

slide-30
SLIDE 30

TID Quantification of Essential Oils

Standard Mix

Analyte Mass % Analyte Mass % α-Pinene 5.15 Isopulegol 4.73 Terpinolene 5.03 Linalool 4.72 β-Pinene 5.01 Geraniol 4.67 γ-Terpinene 4.91 cis-Ocimene 4.66 Camphene 4.90 β-Caryophyllene 4.65 cis-Nerolidol 4.83 α-Humulene 4.65 Limonene 4.83 β-Myrcene 4.64 3-Carene 4.83 trans-Nerolidol 4.58 p-Cymene 4.79 α-Bisabolol 4.53 α-Terpinene 4.77 Guaiol 4.37 trans-Ocimene 4.75

Eucalyptus Essential Oil

Analyte Mass % Analyte Mass % Eucalyptol 68.46 β-Pinene 0.62 α-Terpineol 12.66 Sabinene 0.59 Limonene 4.71 Geraniol 0.58 4-Terpineol 2.69 Linalool 0.37 α-Pinene 2.49 α-Terpinene 0.31 cis-Ocimene 1.37 Terpinolene 0.17 β-Myrcene 1.28 trans-Citral 0.16 α-Phellandrene 1.05 β-Caryophyllene 0.12 trans-Ocimene 0.85 cis-Citral 0.06 p-Cymene 0.80 Nerol 0.03 γ-Terpinene 0.62

slide-31
SLIDE 31

Lavender Essential Oil

Analyte Mass % Analyte Mass % Linalyl acetate 43.72 Camphor 0.31 Linalool 30.67 β-Myrcene 0.30 cis-Ocimene 7.20 Terpinolene 0.24 β-Caryophyllene 3.17 p-Cymene 0.24 4-Terpineol 3.05 γ-Terpinene 0.14 Eucalyptol 2.71 trans-Citral 0.10 trans-Ocimene 2.56 α-Pinene 0.09 α-Terpineol 1.73 β-Pinene 0.08 Geranyl acetate 1.27 Nerol 0.07 Limonene 0.91 Camphene 0.05 Borneol 0.85 Sabinene hydrate 0.02 Caryophyllene oxide 0.53

Neroli Essential Oil

Analyte Mass % Linalool 51.68 Linalyl acetate 8.34 Limonene 7.46 α-Terpineol 7.09 β-Pinene 6.82 trans-Ocimene 4.27 Geraniol 3.63 Geranyl acetate 2.74 trans-Citral 2.25 Nerol 1.98 cis-Ocimene 1.95 trans-Nerolidol 1.51 β-Caryophyllene 0.28

Sweet Orange Essential Oil

Analyte Mass % Limonene 96.02 β-Myrcene 2.32 α-Pinene 0.71 Linalool 0.51 Sabinene 0.31 trans-Citral 0.11 trans-Ocimene 0.02

TID Quantification of Essential Oils

slide-32
SLIDE 32

Peppermint Essential Oil

Analyte Mass % Analyte Mass % Menthol 50.93 α-Terpineol 0.33 Menthone 20.03 α-Terpinene 0.29 Eucalyptol 4.88 Valencene 0.26 Borneol 4.38 β-Myrcene 0.18 Menthyl acetate 4.19 Hydroxycitronellal 0.15 Pulegone 2.80 Terpinolene 0.12 Isomenthone 2.66 4-Terpineol 0.11 β-Caryophyllene 1.77 p-Cymene 0.11 Limonene 1.51 Caryophyllene oxide 0.09 β-Pinene 1.01 trans-Ocimene 0.07 cis-Ocimene 0.94 α-Humulene 0.05 Sabinene hydrate 0.86 Nerol 0.02 α-Pinene 0.76 Citronellol 0.02 Sabinene 0.62 Citronellal 0.02 γ-Terpinene 0.48 Thymol 0.01 Linalool 0.39

Tea Tree Essential Oil

Analyte Mass % Analyte Mass % 4-Terpineol 53.96 Valencene 0.57 γ-Terpinene 17.68 α-Phellandrene 0.48 α-Terpinene 8.29 Limonene 0.38 α-Terpineol 4.51 Caryophyllene oxide 0.21 Terpinolene 3.02 β-Caryophyllene 0.15 p-Cymene 2.37 α-Cedrene 0.10 α-Pinene 2.31 Sabinene 0.10 Eucalyptol 2.21 Linalyl acetate 0.07 cis-Ocimene 2.11 Linalool 0.06 β-Myrcene 0.74 Guaiol 0.04 β-Pinene 0.61 α-Humulene 0.03

TID Quantification of Essential Oils

slide-33
SLIDE 33

Summary of Terpenes Analysis via GC-VUV

  • Terpenes are composed of units of isoprene, which results in a

large isomer prevalence

  • Traditional detection methods such as mass spectrometry cannot

differentiate coeluting isomers; however, because isomers have unique electronic configurations, exposure to VUV light yields distinct spectra

  • GC-VUV works well with multiple sample introduction methods

(liquid injection, headspace, and SPME)

  • 1st
  • rder

absorbance results in linear deconvolution and quantitation

  • f

coeluting analytes; this allows for forced chromatographic compression, leading to shorter run times

  • Potential for automated quantitative analysis using VUV Analyze™
slide-34
SLIDE 34

Thank you for your attention!

And enjoy some terpenes in your favorite adult beverage later!