Advances in Drug Detection in DFSA

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Advances in Drug Detection in DFSA

• Ashraf Mozayani, PharmD.Ph.D, D-ABFT
• Joseph A..Jachimczyk Forensic Center
• Houston, Texas.
• 713-796-6830
• Ashraf_mozayani_at_meo.co.harris.us.tx

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(No Transcript)
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United States

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http//txsdc.tamu.edu/maps/thematic/cnty_totpop.ph
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Harris County
City of Houston
Incorporated Areas
Harris County
Population 3,557,055
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Urine Drug Testing
JAJ Forensic Center Laboratories
Trace
Toxicology
Serology DNA
Controlled Substances
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Overview

• Sexual Assault Statistics
• Definitions
• Signs Symptoms
• Drugs Used
• Specimen Collection
• Toxicological Analysis

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Definitions

• Sexual AssaultForced, coerced or pressured
  sexual contact
• Acquaintance RapeNonconsensual sexual contact
  with someone who is known to you
• Date RapeA type of acquaintance rape, whereby
  nonconsensual sex takes place between two people
  in a relationship
• Drug-Facilitated Sexual AssaultUse of a chemical
  agent to procure sexual contact

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Signs of DFSA

• Confusion
• Dizziness
• Drowsiness
• Psychomotor impairment
• Impaired judgment
• Reduced inhibitions
• Slurred speech

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Signs of DFSA

• If you recall having a drink but cannot recall
  what happened for a period of time afterwards
• If you suspect someone had sex with you but you
  cannot remember any or all of the incident
• If you feel more intoxicated that your usual
  response
• If you wake up feeling hung over and experience
  memory loss or cant account for a period of
  time

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Problems

• Reduced ability to ward off attacker
• Impaired judgment
• Memory loss - anterograde amnesia(drug
  temporarily disables ability to store information
  into memory)
• Victim uncertain of facts
• Unwilling to report the assault
• Depressant-type effects of drugs used to
  incapacitate victim may be very similar to
  alcohol

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Chemical Submission

• Any drug that has the ability to render the
  victim
• Passive
• Submissive
• Unwilling to resist
• Unable to resist

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Chemical Submission Agents

• Potent, fast-acting drugs, some with amnesic
  properties
• CNS depressants that impair consciousness,
  memory, lower inhibitions
• Anesthetic-type effect, unarousable sleep, out
  of body experience, powerless, paralyzed, unable
  to move

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Chemical Submission Agents

• Commonly reported date rape drugs by the
  mediaRohypnol? (Flunitrazepam, Roofies)GHB
• Prevalence of date rape drugs in actual cases of
  alleged sexual assault shows this NOT to be the
  case

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DFSA FACT

• More than 20 substances have been associated with
  drug-facilitated sexual assault
• Benzodiazepines Barbiturates Antihistamines
• Muscle relaxants Stimulants Depressants
• Tranquilizers Narcotics Antidepressants
• Psychedelics Hypnotics Anticonvulsants

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Drugs Associated With Sexual Assault

• Alprazolam
• Amphetamine
• Barbiturates
• Carisoprodol
• Chloral hydrate
• Chlordiazepoxide
• Clonazepam
• Cocaine
• Cyclobenzaprine
• Diazepam
• Diphenhydramine
• Ethanol
• Flurazepam

• Flunitrazepam
• GHB
• Ketamine
• Lorazepam
• Marijuana
• Meprobamate
• Methamphetamine
• Methylenedioxymethamphetamine
• Opiates
• Scopolamine
• Triazolam
• Source J. Forensic Sci. 44 227 (1999)

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Prevalence of Drugs Used in Alleged Cases of
Sexual Assault
J. Anal. Toxicol. 23, 141 (1999)
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Prevalence of Drugs Used in Alleged Cases of
Sexual Assault

• 1179 urine samples, 49 states, 26 months
• Alcohol 41
• THC 19
• Benzos 8
• Cocaine 8

• Amphetamines 4(12.5 in CA)
• GHB 4(8 in CA)
• Opiates 2
• Barbiturates 1
• Propoxyphene 1

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Benzodiazepines

• Total of 97 confirmed positive BZDs
• 72 diazepam, nordiazepam, oxazepam
• 12 Lorazepam
• 8 Alprazolam metabolites
• 6 Flunitrazepam metabolites
• 4 Temazepam

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Multiple Drug Use

• 35 of samples were positive for multiple drugs
• Common combinations
• Alcohol and THC/cocaine/benzodiazepinesAmphetamin
  es and THC/benzodiazepinesGHB and
  alcohol/benzodiazepines/THCGHB alcohol -
  potentiating effect-Increases risk of profound
  sedation, coma

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Caveats

• Analytical factors may result in
  under-representation of certain drugs e.g. GHB,
  low dose benzodiazepines
• Presence of the substance in the urine does not
  necessarily indicate involvement of the drug
  during the incident

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Why have GHB Rohypnol? attracted so much
attention?

• Rapid onset
• Rapid biotransformation
• Short detection windows
• Detection difficulties
• Easily disguised

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Recommendations for Toxicologists

• Develop a routine
• Know your immunoassays
• Improve the detection limits of your analytical
  procedures
• Know your limitations
• Promote research to better understand DFSA

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Developing a Routine for the Non-Routine

• Without a plan, many of these cases get batched
  in with routine cases
• Miss the extra attention that may be necessary
• The lab should develop a routine to address these
  cases

• Cases to accept
• Specimens required and acceptable time to
  collect specimens after ingestion
• What information is needed
• How specimen will be treated to improve
  sensitivity
• Which drugs they will and will not routinely
  screen for
• When to call it quits
• Reporting results

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Good DFSA Cases

• Victim reports immediately
• Provides blood and urine specimen for
  toxicological analysis
• Thorough investigation of suspect that uncovers
  drug(s) likely used
• Analysis can target specific drug(s) and
  metabolite(s)

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Typical DFSA Case

• Victim reports 24-72 hrs after ingestion
• Only blood is collected for DNA purposes
• Investigation waits for toxicology lab to prove
  a drugging occurred

• Labs should develop criteria to determine which
  cases they accept for analysis and which to
  refuse when their analyses will not provide
  probative information
• Specimens
• Information

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Specimens for DFSA Investigations

• Urine
• Collected within 96 hrs of suspected ingestion of
  an agent
• No preservative, but refrigerated or frozen
• At least 100 mL

• Blood
• Collected within 24 hrs of suspected ingestion of
  agent
• Sodium fluoride and potassium oxalate as
  preservatives
• About 10 mL of blood in addition to the urine

M. LeBeau, et al. J. Forensic Sci. 1999, 44(1)
227-230.
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Specimens for DFSA Investigations

• Hair
• Still questionable
• May prove useful for some DFSA drugs, but a
  single dose of a low-dose sedative may not be
  detectable
• Sweat
• Good idea
• Put patch on victim for a few days to a week
• More research needed to determine practicality
  and sensitivity
• Vomit
• May have a high concentration of unabsorbed drug

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Sample Questions to for DFSA Investigators

• What symptoms did the victim describe?
• How long was the victim unconscious?
• What specimens were collected?
• How much time passed between the alleged drugging
  and the collection of specimens?
• How much ethanol did the victim consume?
• Did the victim take any drugs (recreational,
  prescription, or OTC)?
• Did the victim urinate prior to the collection of
  a urine sample? If so, how many times?
• What drugs does the suspect have available to
  use?

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Immunoassays
Y
Y
Y
Signal
Conc
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Must Get Over the Bar
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Immunoassays

• Cross-Reactivity for all the drugs within a
  particular class is not equal
• Some give much better responses than others
• Particular problem with the benzodiazepines
• Improvements seen with techniques such as ELISA
• LOOK at the data, dont let the computer make the
  call
• Look for rises from the baseline set by negative
  controls
• Consider additional, more sensitive assays to
  verify a negative result
• Know the limitations of the immunoassay used in
  your laboratory

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Improving Sensitivity of Established SOPs

• Increase specimen volume to improve chances of
  analyte detection
• Hydrolyze urine specimens for benzodiazepine
  metabolites

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Acid vs Enzymatic Hydrolysis

• Acid
• Rapid and Easy
• Can lower LOD
• Dirty
• Results in benzophenone formation
• Confusion between benzodiazepines
• ID of benzodiazepine?

• Enzymatic
• More Time-Consuming
• Requires enzyme
• Most, but not all
• Cleaner
• Frees the parent benzodiazepine for identification

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Improving Sensitivity of Established SOPs

• Increase specimen volume to improve chances of
  analyte detection
• Hydrolyze urine specimens for benzodiazepine
  metabolites
• Derivatives
• GC/MS(SIM) analysis
• GC/MS(CI) analysis
• New technology

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Flunitrazepam and Metabolites by LC/MS(SIM)
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Confirmation of 7-Aminoflunitrazepam by LC/MS/MS
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Know Your Limitations

• DONT let pride hamper an investigation
• DO let pride push you into improving and
  validating your analytical methods
• Establish a list of routine drugs that your lab
  can detect at subtherapeutic levels
• This list should differ from one lab to the next
• If your caseload does not allow you to direct the
  appropriate attention to the case, then recommend
  another lab do the testing

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Sample List of Routine DFSA Drugs

• Ethanol
• GHB, GBL, 1,4-BD
• Benzodiazepines

• Alprazolam
• Chlordiazepoxide
• Clonazepam
• Diazepam
• Flunitrazepam
• Lorazepam
• Temazepam
• Triazolam

• Amphetamines
• Cocaine
• Marijuana
• Opiates
• Barbiturates
• Sedative Antidepressants
• Sedative Antihistamines

LODs must be low
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Sample List of Non-Routine DFSA Drugs

• Chloral hydrate
• Ethchlorvynol
• Valproic acid
• Dextromethorphan
• p-Methoxyamphetamine
• Scopolamine
• Ketamine
• Tiletamine

• Carisoprodol
• Cyclobenzaprine
• Meprobamate
• Clonidine
• Zolazepam
• Zolpidem

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Promoting Research

• Develop improved immunoassays
• Explore the analysis of alternative specimens
• Hair, Saliva, Sweat
• Synergistic effects of EtOH DFR drugs
• Determine baseline level of GHB in humans

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Calling it Quits

• Specimen volume
• Cost
• Professional judgment of Toxicologist

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Conclusion

• Improvements in drug detection in DFSA cases
  depend on a number of factors
• Laboratories should consider establishing a
  protocol for addressing these cases
• Laboratories should recognize and respect the
  limitations of their immunoassays, as well as
  their other procedures
• Laboratories should strive to improve the
  sensitivity of their standard procedures used for
  these drugs
• Laboratories should promote research in this area

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Conclusions

• DFSA cases are increasing
• Many different drugs are being used - not just
  the ones reported in the media
• Many of the challenges of DFSA can be overcome
• Education is key
• A thorough investigation is a must
• Teamwork is vital

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Drug-Facilitated Sexual Assault A Forensic
Handbook

• The Victim - Abarbanel
• The Perpetrators and Their Modus Operandi -
  Welner
• Ethanol - Garriott and Mozayani
• Rohypnol and Other Benzodiazepines - Robertson
  and Raymon
• Gamma-Hydroxybutyrate (GHB) and Related Products
  - Ferrara, Frison, Tedeschi and LeBeau
• Hallucinogens - Raymon and Robertson
• Opioids - Jufer and Jenkins
• Miscellaneous Prescription and Over-the-Counter
  Medications - Jones and Singer
• Collection of Evidence from DFSA - LeBeau and
  Mozayani
• Analysis of Biological Evidence from DFSA Cases -
  LeBeau and Noziglia
• Sexual Assault Nurse Examiners - Ledray
• Investigating DFSA Cases - Archambault, Porrata
  and Sturman
• Prosecution of DFSA - Kerlin, Riveira and
  Paterson