Barry K Logan PhD, DABFTDirector of Forensic Services

IACP DRE Conference 2010Pittsburgh PA

New Highs: Salvia and K2:

Solutions for the DRE Program

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• Blood/Urine• Scope: Cannabinoids, Cocaine

metabolite, Amphetamines, MDMA, PCP, Barbiturates, Benzodiazepines, Opiates, Methadone, Oxycodone, Propoxyphene.

• Confirmations included in price.• Proactive contact on negative cases.

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Synthetic Cannabinoids

Synthetic drugs that mimic the effects of cannabis.

Investigational use for appetite, blood pressure, nausea, etc

First synthesized in the 1970’s

Sprayed onto dried plant leaves, flowers and stems, and smoked.

Legal or unregulated in most US states.

THCHU-210

CP 47,497

Marijuana to Spice

Cannabicyclohexanol

JWH-018 JWH-073

Clemson Series - JWH

JWH-250

Cannabinoid Mechanism of ActionCerebral Cortex

Higher Cognitive FunctionBasal GangliaCerebellum

MovementHippocampus

Learning, Memory, StressHypothalamus

AppetiteSpinal Chord

Pain, Peripheral sensationMedulla

Nausea, Vomiting

Herbals - Botanical Substrate

Glycyrrhiza glabra(Cultivated Liquorice)

Astragalus membranaceus(Milk Vetch)

Verbascum thapsus(Common Mullein)

(Uchiyama et al, 2010)

Synthetic Cannabinoids

Synthetic Cannabinoids – Drug ID• Agilent 5973 Mass Spectrometer• Agilent 6890 Gas Chromatograph• Electron Impact mode• Full scan• Total run time 10 minutes• Sensitivity 20-50 ug/g

Synthetic Cannabinoids – Drug ID

JWH-018 JWH-073 JWH-200HU-210/11JWH-019JWH-200

WIN 55,212 JWH-015JWH-133 JWH-250 CP55,940

CP 47,497 (n=7) CP 47,497 (n=8)

K2 Blends - Drug Content

(* Uchiyama et al, 2010)

JWH-018 (mg/g)

JWH-073 (mg/g)

CP47,497 (n=8) (mg/g)

JWH-250 (mg/g)

K2 Blonde 12 13 - -K2 Standard 9 9 - -

K2 Citron 10 10 - -K2* (Unknown) - - 6 -

K2 Summit 11 9 - -K2 Blue 15 - - -K2 Pink 11 - - -K2 Latte 16 0.28 - 14K2 Mint 19 0.30 - -K2 Silver 8 - - 16

Spike Gold 20 11 - -Spike Maxx 17 - - 19

Spike Diamond 17 0.07 - -Spike Silver 9 16 - -

Space 10 - - -Herbal blends* 2.0 – 35.9 - 1.1 – 16.9 -

Missouri K2 Administration Study

•IRB Human subjects approval obtained from University of

Central Missouri.

•Six subjects smoked K2 Summit, Citron, Standard

•Each contained JWH-018 and JWH-073, or CP47,497

•Subjects performed SFST’s, cognitive tests and DRE Exam.

•Blood, urine and oral fluid collected.

Missouri K2 Administration Study•Onset of effects in about 2-3 minutes

•Dry mouth•Light headedness•Blurred vision•Agitation, Motor restlessness•Time dilation

•DRE Exam•Increased pulse and blood pressure•Lack of convergence•No HGN, or VGN•Pupils normal, muscle tone normal

•SFST’s•3-4 inches of sway, leg body tremors•Loss of balance•Loss of motor coordination

York County PA, DRE Case• Vehicle crashed into snowbank• Subject appeared drunk or high, difficulty speaking, incoherent, mumbling,

slow slurred speech, nonsense, giggling. • Had smoked “Space” less than 2 hours before.• “I’m not a DUI. The stuff I smoked is legal. You can’t arrest me”

• Courtesy Cpl Brian Torkar, PSP

York County PA, DRE Case• Pulse 96-88, BP 150/80• Eyes watery, red, bloodshot, pupils dilated• Eyelids droopy, HGN present, no VGN• Lack of convergence• SFST’s

• 4 inches of sway on Romberg• Lost balance, used arms on WAT• Swayed, used arms on OLS• Poor finger to nose performance

• Courtesy Cpl Brian Torkar, PSP

Synthetic Cannabinoids - Biologicals• Agilent 6230 TOF Mass Spectrometer• Agilent 1200 binary UPLC• ESI source with Nitrogen Jet • High resolution mode.• Total run time 10 minutes• Sensitivity 0.1ng/mL• Confirmed by LCMS/MS

Analyte Accurate Mass

JWH-018 341.45272

JWH-073 327.42584

JWH-250 335.44600

Target analytesMatrix Analytes

BloodSerumOral Fluid(Refrigerate)

Parent Compounds –JWH-018, JWH-073, JWH-200, HU-210, HU-211, JWH-019, JWH-200 WIN 55,212, JWH-015, JWH-133, JWH-250, CP55,940, CP 47,497 (n=7) CP 47,497 (n=8) …

Urine(Freeze)

Ring monohydroxylated metabolites - glucuronidatedRing dihydroxylated metabolites - glucuronidatedDesmethyl ring hydroxylated metabolites - glucuronidated

York County PA, DRE Case• Drug Identification

• “Space” contained JWH-018, traces of JWH-200

• Toxicology• Blood collected approximately 4 hours after use.• Positive for diphenydramine <50ng/mL• Positive for JWH-018 by directed analysis, LC-TOF

K2 use and the DRE Matrix

Drug Symptom Matrix

CNS

Depressant InhalantsDissociative

Drugs CNS

Stimulants Hallucinogens Narcotic

analgesics Cannabis K2

Horizontal nystagmus Yes Yes Yes No No No No No

Vertical Nystagmus Present * Present* Present No No No No No

Lack of Convergence Present Present Present No No No Present Present

Pupil Size Normal Normal Normal Dilated Dilated Constricted Dilated* Normal

Reaction to Light Slow Slow Normal Slow Normal Little to none Normal Normal

Pulse Rate Down Up Up Up Up Down Up Up

Blood Pressure Down Up/Down Up Up Up Down Up Up

Body Temperature Normal Up/Down/Normal up Up Up Down Normal Normal

* High doses

• Salvia; Sal; Magic Mint, Mystic Sage• Ska Pastora; Sally-D; Diviners Sage; Shepherdess's Herb; ska

Maria Pastora; hojas de Maria; yerba de Maria; • “Salvia divinorum is a sprawling perennial herb found in the Sierra

Mazatec region of Mexico. Its leaves contain the extremely potent Salvinorin A. It has a history of buccal use as a divinatory psychedelic, and has been widely available since the mid 1990s primarily as a smoked herb. Its effects are considered unpleasant by many people.”

http://www.erowid.org/plants/salvia/salvia.shtml

Salvia Divinorum

Salvia Divinorum - Use

• Currently sold in head shops, tobacco shops, and online.• SAMHSA estimates 750K – 1M last year use (2007).• “Buy Salvia” Google search generates 285,000 hits.• Available as plants, crushed leaves, extract• Extracts contain Salvinorin A and B

(mg/g) Sal A Sal B

Leaves 3.2 - 5.0 0.10 – 0.17

Extract 4.1 – 38.9

0.26 – 2.42

Tsujikawa et al, 2008

Salvia Divinorum - Effects

• Following smoking, effects are intense but short lived.• Psychedelic–like changes in visual perception.• Mood and somatic changes• Modified perception of external reality.• Decreased ability to interact with self or surroundings

Gonzalez, et al, 2006

Salvia– Adverse Effects

Vohra et al, 2009

Clinical Feature Frequency

Altered Mental state 7

Hallucinations 3

Excessive laughter 1

Dizziness 1

Mydriasis 2

Flushing 3

Diaphoresis 1

Tachycardia 2

Palpitations 1

Hypoglycemia 1

•18 Isolated Salvia

Exposures

Salvia Divinorum - Pharmacodynamics• C11 Salvinorin A administered to baboons (IV).• Extremely rapid uptake• Peak activity achieved in 40 seconds• Represents 3.3% of dose• PET kinetics match onset and dissipation in humans

Hooker et al, 2008

Salvia Divinorum - Metabolism

Tsujikawa et al, 2009

NMS Labs - Salvinorin Analysis• Positive ion electrospray liquid chromatography

tandem mass spectrometry (LC-MS/MS). • Blood, urine, plasma• Single step L/L extraction

Analyte MRM MRM RT (min)

Sal A 433/373 433/295 6.67

Sal A-d3 436/376 N/A 6.67

Sal B 389/313 389/295 5.88

Test requirements available at www.nmslabs.com

NMS Labs – Salvinorin B Analysis

• Sal B• LOQ

– 1ng/mL

• LOD – ~0.25ng/mL

Salvia Divinorum –Human Study

• Two subjects with prior experience with hallucinogens, smoked 75mg of salvia leaves, for three minutes, ad lib.

• Oral fluid collected at 1 hour post smoking.• Urine collected at 0-1.5 hrs, and 1.5-9 hrs.• Sweat patches applied and removed 2 hrs after smoking.

• “Unfortunately… blood collection was not possible… both volunteers experienced intense hallucinations that started about 30 seconds after inhaling peaking at 3-5 minutes, and lasting about 15-20 minutes. During that period blood drawing was considered unsafe, and at the end of the experience the subjects refused blood drawing.”

Picini et al, 2005

Salvinorin A and B – Stability

Days 1 2 9 14 28

Blood NaF 25oC N/Y N/Y N/Y N N

Blood NaF 4oC N/Y N/Y N/Y N N

Blood NaF -20oC Y Y Y Y Y

Plasma NaF 25oC Y Y Y Y N

Plasma NaF 4oC Y Y Y Y Y

Plasma NaF -20oC Y Y Y Y Y

Urine* 25oC Y Y N/Y N N

Urine* 4oC Y Y Y Y Y

Urine* -20oC Y Y Y Y Y

* unpreserved

Acknowledgements

• Analysis• NMS Labs

• Allan Xu• Matt McMullin• Lindsay Reinhold

• Penn. State Police• Cpl Brian Torkar

• Agilent

• Join Forensic Toxicology on www.LinkedIn.com

• Dosing Study• UCMO

• Bob Welsh• Tracey Durbin• DRE’s and

subjects• WSLH

• Amy Miles

NMS Labs Salvia and K2 test codes• Toxicology• 8070B/S/P/U - Drug Impaired Driving/DRE Toxicology

Panel• 4150B/S/P/U - Salvinorin A & B• 7744B/S/P - Special Request for Synthetic Cannabinoids

• Drug Identification• 7201LI/SL - Controlled Substances/Pharmaceuticals• 7472SL - Synthetic Cannabinoids – solid• 7461LI/SL - Salvinorin A identification – liquid/solid