1. Classification of Common California Cannabis Cultivars via Secondary Metabolite Characterization
Mark A. Lewis
President, Napro Research
California, USA

2. Several Years to Achieve Several Goals:
Validated assays & analysis to communicate data
Collect data & valid survey of ‘chemoscape’
Analysis 1: Class cultivars based on chemotype
Analysis 2: Observe intra-genotypic variability
Analysis 3: Asexual vs. hybrid seed comparison
Garbage In =
Out
Raw
Materials
Finished
Products

3. CEA & Unique Opportunity
56 known cultivars
Identified by common name
Sourced/grown by experts
Stress Free Indoor Environments
Organic practices
Soil
1000W HPS
Many cultivars >40 batches tested

4. Stress Free Environment = Good Data
Production Facility Example
Production/Consumer
Interface
Studio A
Studio B
Studio C
Studio G
Studio F
Studio E
Studio D
Meaningful
& Useful
Information
Monitor Grow Conditions and Product Development in Real Time

5. Data Collection – Climate Metrics
Genotype (G) + Environment (E) → Phenotype (P) & (C)
Real Time Alarms

6. Data Collection – Nutrient Metrics
Genotype (G) + Environment (E) → Phenotype (P) & (C)
Precision Feeding

7. Data Collection – Phenotypic MetricsΔ0
Genotype (G) + Environment (E) → Phenotype (P)

8. Data Collection: Chemotype Metrics
Lack of guidance period
Questions that desperately needed answered:
error in sample prep
extract stability
matrix effects
assay performance
RSD’s, RSD’s, RSD’s

9. Raw Terp and Cannab Data
Material and Data Flow
Tier I
Tier II
Extract 1
Extract 2
Sample
Homogenized
Extract 3
Terpenes %
Plant
Mass
Data
Moisture
Data
Mass
Data
Cannabinoids
2.3-36%
Morphological Data
Resource Data
Sample Prep Worksheet
Cannabinoids %
Metabolites B
Metabolites A
Raw Sample Data
Raw Metabolite Data
Raw Metabolite Data
Raw Terp and Cannab Data
Field Reports
Sample Data Archive
Data Analysis
QC Control Charts
MCF Sheets

10. To Name a Few of the 56 Cultivars Examined
Sour Diesel OG Kush Purple Erkle Ken’s GDP Trainwreck Snowcap Dutch Crunch OG Legend Hawaiian
Skywalker OG
Jack Herrer
Old Amsterdam
Pincher’s Creek
Green Candy
Purple Kush
Bubba Kush
Girl Scout Cookies
Big Sky
Blue Dream
Berry White
Blueberry
SFV OG
Congolese
Pineapples Express
Golden Pineapple
Heroijuana
Etc., Etc., Etc.

11. Total Cannabinoid Content Distribution
Max 36.5%, Min 10.2%, AVG 19.9%
The lot-to-lot RSD’s from 1% to 29% with a 12% AVG
Highly sampled cultivars have lot-to-lot variation from 10% to 20%.
Does variability reflect GxE interactions?

12. Total Terpene Content Distribution
Max 2.47%, Min 0.67%, Avg 1.57%
lot-to-lot RSD from 3% to 41%, average of 21%.
Highly sampled cultivars have lot-to-lot variation from 20% to 30%.
Cannabinoid variability < terpenoid variability

13. Cannabinoid-Terpenoid Relationships
Higher EO = higher cannabinoid content (R2 = 0.156)
Only 15% of variability in terpene content explained by cannabinoid content.
85% of variability due to other explanatory variables not included in this analysis.
Pr>F value of <0.0001, thus, correlation real, but very small.
Analysis of the Variance Pr>F

14. 56 genotypes: Total Terp & CB Content

15. Terpene Traits Visualized with COLOR
PhytoPrintTM displays active compounds with a list of their different medicinal uses
© 2015 BHC Group, LLC. All rights reserved. Any unauthorized use of this document or the images or marks above may violate copyright, trademark, and other applicable laws.

16. 41 Terpenoids Analyzed Only presented 17 on PhytoPrintTM
Most ‘relevant’
Intuitively colored, spaced and arranged
Result: Bar color and length allow rapid ID of similar fingerprints as well as minor differences that make each cultivar unique
Bubba Kush
East Coast Sour Diesel
Classic Krypto
The classification system ignores the “minor” terpenes, which can be defining traits of specific genetic lines.
Classic Purps
Blue Dream 2
Old Amsterdam
Confidential
© 2015 BHC Group, LLC. All rights reserved. Any unauthorized use of this document or the images or marks above may violate copyright, trademark, and other applicable laws.

17. PhytoPrintsTM of Cannabis Archetypes
ocimene
terpinolene
limonene
α-pinene & myrcene
α-pinene
Hybrid with two dominant terpenes
myrcene
β-caryophyllene
© 2015 BHC Group, LLC. All rights reserved. Any unauthorized use of this document or the images or marks above may violate copyright, trademark, and other applicable laws.

18. Terpene SuperClasses

19. Indica, Sativa and the Classification System
Indica & sativa designations are confusing for ethnobotanical purposes
Originally species designations
Now there is “mostly sativa”, “mostly indica”, “sativa-like, indica-blah”, etc.
Says little to the consumer about the product they are purchasing
Class designation conveys most relevant information chemotype
LCM1G
Most relevant terpenes (up to three) in a fixed (not ranked) order. Rationale is that organoleptic properties and entourage effects are largely dictated by the major analytes, and small changes in ranking will be difficult to discern.
Chemotype (I, II, III, etc) implies dominant cannabinoid. This is the major driver of the pharmacology.
Final letter represents the second most dominant cannabinoid and indicates “unusual” cannabinoids than can modulate the properties of the first.

20. PCA & Classification Employed relative terpene profiles for clusteringTW OG BD BW
Employed relative terpene profiles for clustering
Different colors = different classes
Lot-to-lot variability does cause some problems.
Some genotypes don’t always land in same class.
Principal component analysis (PCA) and the resulting biplots provide a manner to determine the defining characteristics of a cultivar, without oversimplifying the analysis by ignoring seemingly insignificant traits… Class system ignores minor terpenes, which can be defining traits of specific genetic lines.

21. Single Genotype: Classic OG Kush
MCL1T class
Myrcene, caryophyllene, limonene
>2% terps, >25% CB’s

22. Classic OG Kush: Multiple Classes

23. Classic OG Analysis: Total Oil Content
Baseline
Organic practices + biostimulant
2.3% AVG Terps
24.3% AVG CB’s
Organic practices – biostimulant
1.8% AVG Terps ↓
18.9% AVG CB’s ↓
Conventional + biostimulant
1.5% AVG Terps ↓
28.1 AVG CB’s ↑

24. Variability & Cannabis Quality
Cannabis industry lacks QA/QC
Most producers have yet to establish quality metrics
Hemp only ‘true’ breeding cannabis seed in market place
Unlike corn, you cannot by disease resistant, or bi-color, etc.

25. Genotype Creation: Type II Trainwreck
Trainwreck is TXX1G class
Terpinolene Dominant

26. TXX2T Class Hybrid Cannabinoid Data

27. TXX2T Class Hybrid Terpene Data

28. Terpene SuperClasses
Indica and sativa may be better suited as an ‘effects gauge’ instead of genealogical nomenclature

29. Sample Reports
© 2015 BHC Group, LLC. All rights reserved. Any unauthorized use of this document or the images or marks above may violate copyright, trademark, and other applicable laws.