Amphetamine and Related Drugs

Amphetamine and
Related Drugs
4/27/2009 ١

Narcotics of Natural Origin
-Opium Opium
-Morphine Morphine
-Codeine Codeine
-Thebaine Thebaine
Semi-Synthetic Semi Synthetic Narcotics
-Heroin Heroin
-Hydromorphone
Hydromorphone
-Oxycodone Oxycodone
-Hydrocodone
Hydrocodone
Synthetic Narcotics
-Meperidine Meperidine
-Dextropropoxyphene
Dextropropoxyphene
-Fentanyl Fentanyl
-Pentazocine Pentazocine
-Butorphanol
Butorphanol
Narcotics Treatment Drugs
-Methadone Methadone
-LAAM LAAM
-BuprenorphineNarcotics
BuprenorphineNarcotics
Narcotics
4/27/2009 ٢

Cocaine
Amphetamines
Methcathinone
Methylphenidate
Anorectic Drugs
Khat
Stimulants
4/27/2009 ٣

Depressants
Barbiturates
Benzodiazepines
Flunitrazepam
Gamma Hydroxybutyric Acid
Paraldehyde
Chloral Hydrate
Glutethimide and Methaqualone
Meprobamate
4/27/2009 ٤

Marijuana
Hashish
Hashish Oil
Cannabis
4/27/2009 ٥

Hallucinogens
LSD
Psilocybin & Psilocyn and other Tryptamines
Peyote & Mescaline
New Hallucinogens
MDMA (Ecstasy) and other
Phenethylamines
Phencyclidine and Related Drugs
Ketamine
4/27/2009 ٦

Other Categories
Inhalants like amyl and butyl nitrite, nitrous
oxide and others
Steroids
4/27/2009 ٧

Drug Schedules: Schedule I
• The drug or other substance has a high potential for
abuse.
• The drug or other substance has no currently
accepted medical use in treatment in the United
States.
• There is a lack of accepted safety for use of the drug
or other substance under medical supervision.
• Examples of Schedule I substances include heroin,
lysergic acid diethylamide (LSD), marijuana, and
methaqualone.
methaqualone.
4/27/2009 ٨

Schedule I amphetamine derivatives
2,5-Dimethoxy
2,5 Dimethoxy-4-ethylamphetamine
ethylamphetamine
2,5-Dimethoxyamphetamine
2,5 Dimethoxyamphetamine
3,4,5-Trimethoxyamphetamine
3,4,5 Trimethoxyamphetamine
3,4-Methylenedioxyamphetamine
3,4 Methylenedioxyamphetamine
3,4-Methylenedioxymethamphetamine
3,4 Methylenedioxymethamphetamine
4-Bromo Bromo-2,5 2,5-dimethoxyamphetamine
dimethoxyamphetamine
4-Bromo Bromo-2,5 2,5-dimethoxyphenethylamine
dimethoxyphenethylamine
4-Methoxyamphetamine
Methoxyamphetamine
4-Methyl Methyl-2,5 2,5-dimethoxyamphetamine
dimethoxyamphetamine
5-Methoxy Methoxy-3,4 3,4-methylenedioxyamphetamine
methylenedioxyamphetamine
4/27/2009 ٩

Schedule II
• The drug or other substance has a high potential for
abuse.
• The drug or other substance has a currently accepted
medical use in treatment in the United States or a
currently accepted medical use with severe
restrictions.
• Abuse of the drug or other substance may lead to
severe psychological or physical dependence.
•Examples •Examples
of Schedule II include morphine,
phencyclidine (PCP), cocaine, methadone, and
methamphetamine
4/27/2009 ١٠

Schedule III
• The drug or other substance has less potential for
abuse than the drugs or other substances in schedules
I and II.
• The drug or other substance has a currently accepted
medical use in treatment in the United States.
• Abuse of the drug or other substance may lead to
moderate or low physical dependence or high
psychological dependence.
• Anabolic steroids, codeine and hydrocodone with
aspirin or Tylenol®, Tylenol , and some barbiturates are
examples of Schedule III substances.
4/27/2009 ١١

Schedule IV
• The drug or other substance has a low potential for abuse
relative to the drugs or other substances in Schedule III.
• The drug or other substance has a currently accepted medical
use in treatment in the United States.
• Abuse of the drug or other substance may lead to limited
physical dependence or psychological dependence relative to
the drugs or other substances in Schedule III.
•Examples •Examples
of drugs included in schedule IV are Darvon®, Darvon ,
Talwin®, Talwin , Equanil®, Equanil , Valium®, Valium , and Xanax®.
Xanax
4/27/2009 ١٢

Schedule V
• The drug or other substance has a low potential for
abuse relative to the drugs or other substances in
Schedule IV.
• The drug or other substance has a currently accepted
medical use in treatment in the United States.
• Abuse of the drug or other substances may lead to
limited physical dependence or psychological
dependence relative to the drugs or other substances
in Schedule IV.
•Cough •Cough
medicines with codeine are examples of
Schedule V drugs
4/27/2009 ١٣

Most Common Amphetamines
There are a large number of amphetamines which are
controlled substances. Of these, the most commonly
encountered in the forensic science laboratory are
amphetamine (1), ( ), methylamphetamine (2), ), 3,4- 3,4
methylenedioxyamphetamine (MDA) (3), ( ), 3,4- 3,4
methylenedioxymethylamphetamine (MDMA) (4)and ( )and
3,4-methylenedioxyethylamphetamine 3,4 methylenedioxyethylamphetamine (MDEA) (5). ( ).
In addition, there are a wide variety of structurally
related analogues which can be synthesized.
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4/27/2009 ١٥

Some history
Amphetamine was first marketed in the 1930s as
Benzedrine®
Benzedrine in an over-the over the-counter counter inhaler to treat
nasal congestion. By 1937, amphetamine was
available by prescription in tablet form and was used
in the treatment of the sleeping disorder, narcolepsy,
and the behavioral syndrome called minimal brain
dysfunction, which today is called attention deficit
hyperactivity disorder (ADHD). During World War
II, amphetamine was widely used to keep the fighting
men going and both dextroamphetamine
(Dexedrine®) (Dexedrine ) and methamphetamine (Methedrine ( Methedrine®) )
were readily available.
4/27/2009 ١٦

As use of amphetamines spread, so did their
abuse. In the 1960s, amphetamines became a
perceived remedy for helping truckers to
complete their long routes without falling
asleep, for weight control, for helping athletes
to perform better, and for treating mild
depression. With experience, it became evident
that the dangers of abuse of these drugs
outweighed most of their therapeutic uses.
4/27/2009 ١٧

Increased control measures were initiated in 1965
with amendments to the federal food and drug laws to
curb the black market in amphetamines. Many
pharmaceutical amphetamine products were removed
from the market including all injectable formulations,
and doctors prescribed those that remained less
freely. Amphetamine products presently marketed
include generic and brand name amphetamine
(Adderall Adderall®, , Dexedrine®,
Dexedrine , Dextrostat®) Dextrostat ) and brand
name methamphetamine (Desoxyn ( Desoxyn®). ).
4/27/2009 ١٨

To meet the ever-increasing ever increasing black market demand for
amphetamines, clandestine laboratory production has
mushroomed. Today, most amphetamines distributed
to the black market are produced in clandestine
laboratories. Methamphetamine laboratories are, by
far, the most frequently encountered clandestine
laboratories in the United States. The ease of
clandestine synthesis, combined with tremendous
profits, has resulted in significant availability of illicit
methamphetamine, especially on the West Coast,
where abuse of this drug has increased dramatically
in recent years.
4/27/2009 ١٩

Amphetamines are generally taken orally or
injected. However, the addition of "ice," the
slang name for crystallized methamphetamine
hydrochloride, has promoted smoking as
another mode of administration. Just as
"crack" is smokable cocaine, "ice" is smokable
methamphetamine. Methamphetamine, in all
its forms, is highly addictive and toxic.
4/27/2009 ٢٠

The effects of amphetamines, especially methamphetamine,
are similar to cocaine, but their onset is slower and their
duration is longer. In contrast to cocaine, which is quickly
removed from the brain and is almost completely metabolized,
methamphetamine remains in the central nervous system
longer, and a larger percentage of the drug remains unchanged
in the body, producing prolonged stimulant effects. Chronic
abuse produces a psychosis (severe mental disorder), picking
at the skin, and visual hallucinations. These psychotic
symptoms can persist for months and even years after use of
these drugs has ceased and may be related to their neurotoxic
effects. Violent and erratic behavior is frequently seen among
chronic abusers of amphetamines, especially
methamphetamine.
4/27/2009 ٢١

AMPHETAMINE
Amfetamine
Central Stimulant
Synonyms. Amphetamine; Anfetamina; Racemic
Dexedrine.
Proprietary names. It is an ingredient of
Biphetamine and Durophet.
4/27/2009 ٢٢

A colourless, mobile, slowly volatile liquid. It
absorbs carbon dioxide from the air forming a
volatile carbonate. B.p. 200° 200 to 203°. 203
Soluble 1 in 50 of water; soluble in ethanol
chloroform and ether; readily soluble in acids
Colour Tests.
Liebermann's Test (sulfuric acid + nitrous acid)
—red red–orange; orange; Marquis Test—orange
Test orange→brown; brown;
Ninhydrin—pink
Ninhydrin pink–orange orange
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Amfetamine Phosphate
Synonyms. Amphetamine Phosphate; Benzpropaminum
Phosphoricum; Monobasic Racemic Amphetamine
Phosphate.
FW = 233.2
A white crystalline powder with no characteristic
melting point; it sinters at about 150° 150 and
decomposes at about 300°. 300
Freely soluble in water; slightly soluble in ethanol;
practically insoluble in benzene, chloroform, and
ether.
ether
4/27/2009 ٢٤

Amfetamine Sulfate
Synonyms. Amphetamine Sulfate; Phenaminum; Phenopromini
Sulfas; Phenylaminopropanum Racemicum Sulfuricum;
Psychedrinum.
Proprietary names. Benzedrine; Centramina. It is an ingredient of
Adderall, Epipropane, and Ortenal.
FW = 368.5
A white crystalline powder. M.p. above 300°, 300 , with
decomposition.
Soluble 1 in 9 of water and 1 in 515 of ethanol; practically
insoluble in chloroform and ether.
ether
4/27/2009 ٢٥

Disposition in the Body.
Readily absorbed after oral or rectal administration;
rapidly distributed extravascularly and taken up, to
some extent, by red blood cells. The main metabolic
reaction is oxidative deamination to form
phenylacetone, which is then oxidised to benzoic acid
and conjugated with glycine to form hippuric acid;
minor reactions include aromatic hydroxylation to
form 4–hydroxyamfetamine 4 hydroxyamfetamine (an active metabolite), β-
hydroxylation to form norephedrine
(phenylpropanolamine), and N-oxidation oxidation to form a
hydroxylamine derivative.
4/27/2009 ٢٦

4/27/2009 ٢٧

Excretion of amfetamine is markedly dependent on urinary
pH, being greatly increased in acid urine. After large doses,
amfetamine may be detected in urine for several days. Under
uncontrolled urinary pH conditions, about 30% of the dose is
excreted unchanged in the urine in 24 h and a total of about
90% of the dose is excreted in 3 to 4 days. The amount
excreted unchanged in 24 h may increase to 74% of the dose in
acid urine and decrease to 1 to 4% in alkaline urine; under
alkaline conditions, hippuric acid and benzoic acid account for
about 50% of the urinary material. Under normal conditions 16
to 28% is excreted as hippuric acid, about 4% as
benzoylglucuronide, 2 to 4% as 4–hydroxyamfetamine, 4 hydroxyamfetamine, and
about 2% as norephedrine in 24 h; small amounts of
conjugated 4–hydroxynorephedrine 4 hydroxynorephedrine and phenylacetone are
also excreted. No elimination in the faeces has been detected.
4/27/2009 ٢٨

Therapeutic concentration
After normal therapeutic doses the plasma
concentration is usually below 0.1 mg/L. However,
continued use of amfetamine may cause addiction,
and ingestion of 10 times the usual therapeutic dose is
common among addicts; in such cases the plasma
concentration may be up to 3 mg/L.
After a single oral dose containing 10 mg of
amfetamine to 4 subjects, peak plasma concentrations
of about 0.02 mg/L were attained
4/27/2009 ٢٩

Steady–state Steady state blood concentrations of 2 to
3 mg/L were observed in a regular user who
ingested about 1 g a day.
The intravenous administration of 160 mg of
amfetamine to a regular user resulted in a
plasma concentration of 0.59 mg/L after 1 h.
4/27/2009 ٣٠

Toxicity
The estimated minimum lethal dose in non–addicted
non addicted
adults is 200 mg. Toxic effects may be produced with
blood concentrations of 0.2 to 3 mg/L, and fatalities
with concentrations greater than 0.5 mg/L. Death
from overdosage is comparatively rare.
In a fatality caused by intravenous administration of
amfetamine, the following postmortem tissue
concentrations were reported: blood 41 mg/L, liver
23 μg/g, g/g, urine 39 mg/L.
4/27/2009 ٣١

Half–life. Half life.
Plasma half–life, half life, 4 to 8 h when the urine is acidic and
about 12 h in subjects whose urinary pH values are
uncontrolled.
Volume of distribution.
About 3 to 4 L/kg.
Dose.
20 to 100 mg of amfetamine sulfate daily has been
used in the treatment of narcolepsy.
4/27/2009 ٣٢

Methylenedioxymethamfetamine
Stimulant, Hallucinogen
Synonyms. Adam; Clarity; E; Ecstasy; Elaine;
Essence; Euphoria; MDM; MDMA; 3,4- 3,4
Methylenedioxymetamfetamine; 3,4– 3,4
methylenedioxymethylamphetamine; Stacy; X;
XTC.
4/27/2009 ٣٣

Street names
Several tablets and capsules have been in circulation
over the years, containing varying amounts of
MDMA and other phenethylamines, recognisable by
logos on tablets or colours of capsules. Some
examples of tablets or capsules include: Apples;
Beans; Baby slits; Brownies; Burgers; Crowns;
Dennis the Menaces; Diamond Whites; Disco
biscuits; Dollars; Doves; Love Doves; Mitsubishis;
Mitsies; New Yorkers; Pink Calis; Rhubarb &
Custards; Tangos; White Doves.
4/27/2009 ٣٤

N,α-Dimethyl Dimethyl–1,3,benzodioxole
1,3,benzodioxole–5–
ethanamine
FW =193.2
A viscous, colourless oil. B.p. 100° 100
to 110°. 110 .
4/27/2009 ٣٥

Methylenedioxymethamfetamine
Hydrochloride
FW = 229.75
A white to off–white off white crystalline powder with a
bitter taste. M.p. 147° 147 to 148° 148 for crystals from
isopropanol/n–hexane; isopropanol/ hexane; M.p. 152° 152 to 153° 153 for
crystals from isopropan-2-ol/ether.
isopropan ol/ether.
4/27/2009 ٣٦

Colour Test.
Marquis Reagent—black Reagent black with dark purple.
Thin–layer Thin layer Chromatography.
System TA—Rf TA Rf 0.33; system TB—Rf TB Rf 0.24;
system TE—Rf TE Rf 0.39; system TF—Rf TF Rf 0.20;
system TAE—Rf TAE Rf 0.08; system TAJ—Rf TAJ Rf 0.03;
system TAK—Rf TAK Rf 0.17; system TAL—Rf TAL Rf 0.57.
4/27/2009 ٣٧

Disposition in the Body.
It is absorbed into the blood stream after ingestion and
excreted in urine, the majority of the dose unchanged (65%
within 3 days). Metabolism occurs by a number of routes: N-
demethylation of the parent compound to 3,4– 3,4
methylenedioxyamfetamine (MDA) (7%) with further O-
demethylation to 3,4–dihydroxymethamfetamine 3,4 dihydroxymethamfetamine (HHMA)
and 3,4–dihydroxyamfetamine 3,4 dihydroxyamfetamine (HHA). Both HHMA and
HHA are subsequently O-methylated methylated mainly to 4–hydroxy 4 hydroxy–3–
methoxymetamfetamine (HMMA) and 4–hydroxy 4 hydroxy–3–
methoxyamfetamine (HMA). These four metabolites are
excreted in the urine as the conjugated glucuronide or sulfate
metabolites.
4/27/2009 ٣٨

Therapeutic concentration
8 healthy male volunteers, aged between 21 and 31
years old, were administered a 75 mg dose of
MDMA. The mean peak plasma concentration was
0.13 mg/L after 1.8 h. Mean peak plasma
concentrations of the metabolite, 3,4– 3,4
methylenedioxyamfetamine (MDA), were 7.8 μg/L g/L
approximately 5 h after administration.
4/27/2009 ٣٩

After the administration of a single oral dose of
1.5 mg/kg body weight MDMA to 2 patients, plasma
and urine samples were collected over periods of 9
and 22 h, respectively. Peak plasma concentrations of
MDMA and MDA were 331 μg/L g/L after 2 h and
15 μg/L g/L after 6.3 h, respectively. Peak concentrations
of 28.1 μg/L g/L MDMA in urine appeared after 21.5 h.
Up to 2.3 μg/L g/L MDA, 35.1 μg/L g/L HMMA, and
2.1 μg/L g/L HMA were measured within 16 to 21.5 h,
also in urine.
4/27/2009 ٤٠

Toxicity
Fatalities with doses of 300 mg have been
reported. Capable of causing severe toxicity
and the pattern of acute toxicity is due to the
circumstances in which it is misused. A lethal
concentration of 0.35 to 0.50 mg/L in serum
has been noted although some overdose cases
report concentrations 10 times this amount,
without fatality.
4/27/2009 ٤١

Half–life. Half life.
About 6 to 7 h.
Clearance.
The mean total clearance of MDMA for a 75 mg dose
is 86.9 L/h.
Protein binding
About 65%
Dose.
The usual dose is between 80 and 200 mg (more
often 100 to 150 mg).
4/27/2009 ٤٢

Metamfetamine
Central Stimulant
Synonyms. d-Deoxyephedrine;
Deoxyephedrine;
Desoxyephedrine; Methamphetamine;
Methylamfetamine; methylamphetamine;
Phenylmethylaminopropane.
Note.
Metamfetamine in a smokeable form has been
known as Crank, Crystal, Crystal meth, Ice,
meth, and Speed.
4/27/2009 ٤٣

Proprietary name. name Norodin
FW =149.2
A clear, colourless, slowly volatile, mobile
liquid. Mass per mL 0.921 to 0.922 g. B.p.
about 214°. 214
Slightly soluble in water; miscible with
ethanol, chloroform, and ether.
4/27/2009 ٤٤

Metamfetamine Hydrochloride
Proprietary names. names Amphedroxyn; Desfedrin;
Desoxyfed; Desoxyn; Destim; Doxephrin; Drinalfa;
Gerobit; Hiropon; Isophen; Isophen;
Madrine; Methampex;
Methedrine; Methylisomyn; Pervitin;
Soxysympamine; Syndrox; Tonedron.
FW =185.7
White crystals or crystalline powder. M.p. 170° 170 to
175°. 175
Soluble 1 in 2 of water, 1 in 4 of ethanol, and 1 in 5
of chloroform; practically insoluble in ether.
4/27/2009 ٤٥

Colour Test.
Marquis Test—orange.
Test orange.
Thin–layer Thin layer Chromatography.
System TA—Rf TA Rf 0.31; system TB—Rf TB Rf 0.28; system
TC—Rf TC Rf 13; system TE—Rf TE Rf 0.42; system TL—Rf TL Rf
0.05; system TAE—Rf TAE Rf 0.09; system TAF—Rf TAF Rf 0.63;
system TAJ—Rf TAJ Rf 0.00; system TAK—Rf TAK Rf 0.03; system
TAL—Rf TAL Rf 0.45. (Dragendorff spray, positive;
acidified iodoplatinate solution, positive; Marquis
reagent, brown; ninhydrin spray, positive; acidified
potassium permanganate solution, positive.)
4/27/2009 ٤٦

Disposition in the Body
Readily absorbed after oral administration. About
70% of a dose is excreted in the urine in 24 h. Under
normal conditions, up to 43% of a dose is excreted as
unchanged drug, up to 15% as 4– 4
hydroxymetamfetamine, and about 5% as
amfetamine, amfetamine,
the major active metabolite. A number
of other metabolites have been identified. Excretion
of unchanged drug is dependent on the urinary pH,
being increased in acidic urine and greatly reduced
(to about 2% of a dose) if the urine is alkaline
4/27/2009 ٤٧

Following a single oral dose of 12.5 mg of
metamfetamine hydrochloride to 10 subjects, a
mean peak blood concentration of about
0.02 mg/L was attained in about 2 h
Toxicity
The estimated minimum lethal dose is 1 g, but
fatalities attributed to metamfetamine are rare.
4/27/2009 ٤٨

Half–life Half life
Plasma half–life, half life, about 9 h.
Dose.
2.5 to 25 mg of metamfetamine hydrochloride
daily, by mouth; 15 to 20 mg IM, or 10 to
15 mg IV.
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Methylenedioxyethylamfetamine
Stimulant, Hallucinogen
Synonyms. N-Ethyl Ethyl–3,4 3,4–
methylenedioxyphenylisopropylamine; Eve; MDE;
MDEA; 3,4-Methylenedioxyethamphetamine; 3,4 Methylenedioxyethamphetamine; 3,4- 3,4
Methylenedioxyethylamphetamine. Usually presented
as Ecstasy.
N-ethyl ethyl-α-methyl methyl–1,3 1,3–benzodioxole
benzodioxole–5–ethanamine ethanamine
FW =207.3
4/27/2009 ٥٠

A viscous, colourless oil. B.p. is 0.2 is 0.2
85° 85 to 95°
95
4/27/2009 ٥١

Methylenedioxyethylamfetamine
Hydrochloride
FW = 243.8
A white to off–white off white crystalline powder with a
bitter taste.
4/27/2009 ٥٢

Disposition in the Body.
It is absorbed into the blood stream after
ingestion and excreted in urine, mainly as the
parent drug (19%),
methylenedioxyamfetamine (MDA, 28%) and
also 4–hydroxy 4 hydroxy–3–methoxyethylamfetamine
methoxyethylamfetamine
(HMEA, 32%).
4/27/2009 ٥٣

Toxicity
The estimated lethal dose is 0.5 g.
In a 20-year 20 year-old old male whose death was
attributed to injection of MDMA and MDEA,
postmortem blood concentrations of 2.0 and
0.7 mg/L, respectively, were reported
4/27/2009 ٥٤

Methylenedioxyamfetamine
Hallucinogen
Synonyms. MDA;
Methylenedioxyamphetamine;
Tenamfetamine; SKF-5. SKF 5.
α-Methyl Methyl–1,3 1,3–benzodioxole
benzodioxole–5–ethanamine ethanamine
FW =179.2
4/27/2009 ٥٥

4/27/2009 ٥٦

Why are adulterants and diluants
added to a sample?
There are a number of reasons for these additions.
Adulterants, for example, caffeine and other
pharmacologically active drugs, are added to either
hide the lack of the desired drug, dilute it (and hence
increase profit pro for the drug dealer), or add another
type of effect to the drug mixture. Diluents are
bulking agents and may include starch, talc, etc., and
are added to make the drug ‘go go further’. further . Some
additives may be added to the powders, either to
improve the flow ow properties of the tablets prior to the
tableting process, or to impart a particular color.
color
4/27/2009 ٥٧

Qualitative Identification
Identi cation of
Amphetamines
A wide variety of types of sample may be
expected by the forensic scientist. These
include powdered drug material, tableted
material and items likely to have traces of the
drug samples present. From these items, the
appropriate samples should be taken, described
and subsequently forwarded for analysis .
4/27/2009 ٥٨

Sampling and Physical Description of
Amphetamines
Powder Samples
If the samples are powdered materials, they should be sorted
into groups, where the members of the groups cannot be
distinguished from each other. Having achieved this, the items
in each group should be counted and a good physical
description prepared. This should include weight, color, odor
and any other physical characteristic that the scientist
considers to be important. Depending upon the number of
items in the group, the following sampling strategy, based
upon the United Nations Drug Control Program (UNDCP)
recommendations, may be adopted. If there are between 1 and
10 items, all of them should be examined. If there are between
10 and 100 items, then 10 items should be examined, while if
there are more than 100 items, the square route of the number
of items should be examined
4/27/2009 ٥٩

Tableted Samples
If the items are tableted, tableted,
then a different approach is required.
The items should be divided into visually indistinguishable
groups and the number of items in each group should be
counted, if necessary estimating the number from the mean
weight of tablet and the total mass of the tablets, if very large large
numbers are involved. A good physical description of the
tablets should be made, including recording of the size, color,
shape, logo and score marks (if present), while the ballistics
(physical characteristics) of the tablet should be examined and
detailed. Photography is particularly helpful in this respect.
This latter includes recording of all of the physical damage to
the tablets which may be present.
4/27/2009 ٦٠

Recording the ballistic features of tableted
drug units
Having recorded all of the physical data, the items to
be examined chemically should be chosen. At the
time of writing, there is no agreed best-practice
best practice
protocol for undertaking this and the analyst should
work within the requirements of the judicial system in
which he/she is operating. The theory applied to
powdered amphetamines can be equally applied to
amphetamine tablets. The latter should be sampled
and presumptive tests, TLC and confirmatory
con rmatory tests
then carried out.
4/27/2009 ٦١

If the samples are trace samples, that is, the drug
present is likely to be easily contaminated, the
approach should be that the operators, laboratory
equipment and reagents to be used should be
demonstrably free of drug residues prior to the
commencement of the analysis. This can be achieved
by washing the glassware, work surfaces and
operator’s operator s hands with a small amount of methanol and
concentrating the dissolved materials. The same batch
of solvent should be used for this procedure and for
the analysis of the drug items themselves
4/27/2009 ٦٢

Having carried out the control procedures, the item(s) item(s)
to be examined should then be swabbed, individually,
by using a clean swab soaked in a suitable solvent. A
suitable solvent should freely dissolve the drug
material, not cause decomposition of the drug or react
with it, and be amenable to subsequent analytical
procedures. It should be remembered that the
swabbing process should leave enough material intact
for a second and subsequent analysis. If the swab is
not to be used immediately, it should be dried and
stored until needed.
4/27/2009 ٦٣

Sample Homogenization
Homogenization of powdered samples can be achieved by
using a number of different methodologies. One of the best
methods for samples likely to be encountered ‘on on the street’ street is
the use of the ‘cone cone-and and-square square’ method. In this technique,
the materials are mixed and the larger fragments reduced in
size. The material is poured onto a flat at, , clean surface and then
divided into four; opposite quarters are removed and the two
remaining quarters recombined. The process is repeated until
the desired sample size is achieved. For larger samples, a ‘core core’
may be taken and then subjected to further homogenization by
using this cone-and cone and-square square methodology.
methodology
4/27/2009 ٦٤

For tableted materials, homogenization is more
problematic since if the tablet was
homogenized, it would also be destroyed. For
this reason, a sample is taken from the tablet
and gently scraped from the dose form, away
from any ballistic features which are likely to
be of use in subsequent examinations. The
powder is then thoroughly homogenized prior
to testing .
4/27/2009 ٦٥

Thin Layer Chromatography of
Amphetamines
In order that the sample can be tested for the presence
of amphetamines, a test solution must be prepared.
The sample should be dissolved in a suitable solvent
(methanol is commonly used) at a sample
concentration of the order of 10 mgml −1 . This allows
for the fact that many amphetamine samples at the
‘street street level’ level are extremely weak, i.e. between 2 and
10% amphetamine in a matrix of adulterants and
diluants, diluants,
giving a solution of approximately 0.2–1.0 0.2 1.0
mgml −1 , namely a concentration at which the
standards can be prepared.
4/27/2009 ٦٦

The sample should be dissolved as fully as
possible and centrifuged or filtered ltered to remove
any solid particulates. A positive and negative
control should also be prepared. The silica gel
chromatographic plate should be marked up
and the test solutions, plus the positive and
negative controls, placed on the plate and the
latter allowed to develop in the chosen solvent
system
4/27/2009 ٦٧

Practical TLC Urine Tests
The urine sample is not hydrolyzed for Test B. The
urine sample is adjusted to pH 10 with potassium
carbonate. Sodium chloride is also added and the
mixture is extracted twice with chloroform. The
chloroform phase each time is removed and filtered.
The pooled chloroform extracts are washed with a
weak solution of ammonium hydroxide. The washed
chloroform is then extracted twice with 1 N sulfuric
acid.
acid
4/27/2009 ٦٨

The pooled sulfuric acid extracts are then
adjusted to pH 10 with concentrated potassium
hydroxide and potassium carbonate. Sodium
chloride is also added and the mixture is
extracted twice with chloroform. The filtered
and pooled chloroform is then carefully
evaporated after the addition of one drop of a
solution of 0.5% sulfuric acid in methanol.
4/27/2009 ٦٩

The residue is redissolved in acetone methanol
solution and applied to a T.L.C. plate for
development. Test B solvent system contains
methanol and ammonium hydoxide. hydoxide.
Test B
detects methadone, pethidine, pethidine,
cocaine,
amphetamine, methamphetamine, cyclazocine
and D-propoxyphene
D propoxyphene. . Many other organic
bases would be extracted by this procedure and
appear on the T.L.C. plate.
4/27/2009 ٧٠

Urine analyzed by Test B shows an increase in
background with the age of urine which partially
interferes with the location of the spots after T.L.C.
Amphetamine, methamphetamine, pethidine and
methadone are more labile compounds than morphine
and codeine and more susceptible to decomposition
and chemical change during storage or during testing.
testing
4/27/2009 ٧١

Procedure for Test B
Measure 20 ml of urine into a 50 ml glass- glass
stoppered centrifuhe tube. Add 1 g of
potassium carbonate to adjust the pH to 10.
Add 4 g of sodium chloride. Add the salts
using a powder funnel and measuring spoons.
Shake to dissolve the salts.
Add 20 ml of chloroform and shake for 5
minutes and centrifuge.
4/27/2009 ٧٢

Aspirate off the lower chloroform layer and filter
into another tube.
Add 20 ml of chloroform for a second extraction.
Shake for 5 minutes and centrifuge.
Aspirate off the lower chloroform layer and filter
into the second tube.
Wash the filtered pooled chloroform with 10 ml of
pH 9 aqueous ammonium hydroxide solution as
follows: shake for 5 minutes and centrifuge and
aspirate off and discard the upper wash phase.
phase
4/27/2009 ٧٣

Add 10 ml of 1 N sulfuric acid to the tube, shake for
5 minutes and centrifuge. Aspirate off the upper
acid phase and tranfer it to a third tube.
Repeat the extraction with another 10 ml portion of
1 N sulfuric acid and pool with the first extraction
in the third tube. Discard the lower chloroform
phase.
To the acid phase in the third tube add 16 N
potassium hydroxide dropwise (about 1.3 ml) to
adjust the pH to about 7. Add 1 g of potassium
carbonate to adjust the pH to 10. Add 4 g of sodium
chlorides and shake to dissolve the salts.
4/27/2009 ٧٤

Add 20 ml chloroform and shake for 5 minutes and
centrifuge. Aspirate the lower chloroform phase and
filter into a 50 ml beaker.
Repeat the extraction with a second 20 ml of
chloroform and aspirate off and discard the upper
aqueous phase. Decant and filter the chloroform
phase into the beaker. Add 3 ml of chloroform wash
to the tube and filter into the beaker.
4/27/2009 ٧٥

Add one drop of 0.5 % sulfuric acid in
methanol to the pooled chloroform in the
beaker.
Evaporate carefully to near dryness in the
vacuum oven at a temperature of 90 oC and a
vacuum of 10 p.s.i. p.s.i.
Remove the beaker and
allow the final few drops of solvent to air
dry.
4/27/2009 ٧٦

Transfer the residue to a 3 ml
microcentrifuge tube using small portion (0.5
ml) of 1:1 acetone-methanol. acetone methanol. Again
evaporate at a slow boil to near dryness in
the vacuum oven maintained at 10 p.s.i, p.s.i,
and
60 oC. 4/27/2009 ٧٧

Remove a thin-layer thin layer plate from the desiccator
just before it is to be spotted. Spot the sample
residues, procedure controls and reference
compounds on a thin-layer thin layer plate on the sample
application line located 2.5 cm and parallel to
the bottom edge of the plate.
4/27/2009 ٧٨

Dissolve the residues in 20 µl l of 1:1 acetone- acetone
methanol and spot the dissolved sample from
the micro centrifuge tubes using a 10 µl l
microsyringe. microsyringe.
Repeat the spotting twice adding
solvent each time to insure that all of the
dissolved residue is transferred. Apply 30µg 30 g of
methadone, 60 µg g each of amphetamine and
methamphetamine toward the center of the
application line .
4/27/2009 ٧٩

Place the spotted plated into the developing
tank containing 3 ml of conc. ammonium
hydroxide in 200 ml of methanol which has
equilibrated for 10 minutes. Allow the
development to proceed until there is about
14 cm of front movement in about 30
minutes.
4/27/2009 ٨٠

Remove the developed plate and allow it to air
dry for about one hour. Examine the plate
under ultraviolet light for absorbing or
fluorescent spots and circle the spots on the
uncoated side of the plate using a china
marking pencil. Spray with Dragendorff's
Reagent and make notes of spots, colors and
intensities. Then spray with potassium
iodoplatinate reagent and repeat the
observations .
4/27/2009 ٨١

Examine the batch of plates making
appropriate comparisons. Spray the plates
within two hours after development and read
and interpret them as the sprays are applied
and again 10 minutes later.
4/27/2009 ٨٢

Methamphetamine (Methedrine ( Methedrine) ) 44(B)
Amphetamine (Benzedrine) 54(B)
Solvent System for Test B - Mixture of 200 ml
of methanol and 3 ml of concentrated
ammonium hydroxide.
4/27/2009 ٨٣

The ammonia is added to achieve a process known as
ion suppression. suppression.
By converting the drugs to their
free base forms, their polarities are reduced. This is
because the nitrogen atom does not carry a positive
charge in basic solution. The latter reduces the
problem of (TLC) tailing, improves the mass transfer
properties between the stationary and mobile phases,
and thus improves the chromatographic quality.
4/27/2009 ٨٤

In addition, MDA, MDMA and MDEA give rise to
purple, orange/red and orange/red products,
respectively. At each of the visualization stages, the
retardation factor (or relative front) (Rf ( Rf) ) values of the
visualized compounds should be calculated by using
the following equation:
Distance moved by the analyte of interest
Rf = -------------------------------------------------------
Distance moved by the solvent front
4/27/2009 ٨٥

The Rf values of the unknowns are compared to those
of the standards and if the data cannot be
discriminated then a suggested match is called.
Although when using this combination of
presumptive tests and TLC it is possible to
discriminate within this group of compounds, due to
the extremely large number of amphetamines
available, it is necessary to carry out a confirmatory
con rmatory
analytical technique. The foremost of these, for
amphetamine identification
identi cation, , is gas chromatography–
chromatography
mass spectrometry (GC–MS (GC MS)
4/27/2009 ٨٦

Thin–layer Thin layer Chromatographic Systems
for Amphetamine
System TA—Rf TA Rf 43; system TB—Rf TB Rf 20;
system TC—Rf TC Rf 09; system TE—Rf TE Rf 43; system
TL—Rf TL Rf 18; system TAE—Rf TAE Rf 12; system
TAF—Rf TAF Rf 75. (Dragendorff spray, positive;
FPN reagent, pink; acidified iodoplatinate
solution, positive; Marquis reagent, brown;
ninhydrin spray, positive; acidified potassium
permanganate solution, positive). positive).
4/27/2009 ٨٧

System TA
Plates: Plates:
Silica gel G, 250 μm m thick, dipped in,
or sprayed with, 0.1 M potassium hydroxide in
methanol, and dried.
Mobile phase: phase:
Methanol:strong ammonia
solution (100:1.5).
Reference compounds: compounds:
Atropine Rf 18,
Codeine Rf 33, Chlorprothixene Rf 56,
Diazepam Rf 75.
4/27/2009 ٨٨

Colour test
The Marquis test gives an orange colour for both
amfetamine and metamfetamine.
Thin layer chromatography
TA: amfetamine Rf = 0.43, metamfetamine Rf = 0.31.
TB: amfetamine Rf = 0.15, metamfetamine Rf = 0.28.
Visualisation: acidified iodoplatinate solution.
4/27/2009 ٨٩

System TB
Plates: Plates:
Silica gel G, 250 μm m thick, dipped in,
or sprayed with, 0.1 M potassium hydroxide in
methanol, and dried.
Mobile phase: phase:
Cyclohexane:toluene
Cyclohexane: toluene:diethylamine
diethylamine (75:15:10).
Reference compounds: compounds:
Codeine Rf 06,
Desipramine Rf 20, Prazepam Rf 36,
Trimipramine Rf 62
4/27/2009 ٩٠

System TC
Plates: Plates:
Silica gel G, 250 μm m thick, dipped in,
or sprayed with, 0.1 M potassium hydroxide in
methanol, and dried.
Mobile phase: phase:
Chloroform:methanol (90:10).
Reference compounds: compounds:
Desipramine Rf 11,
Physostigmine Rf 36, Trimipramine Rf 54,
Lidocaine Rf 71.
4/27/2009 ٩١

System TL
Plates: Plates:
Silica gel G, 250 μm m thick, dipped in,
or sprayed with, 0.1 M potassium hydroxide in
methanol, and dried.
Mobile phase: phase:
Acetone.
Reference compounds: compounds:
Amitriptyline Rf 15,
Procaine Rf 30, Papaverine Rf 47, Cinnarizine
Rf 65.
4/27/2009 ٩٢

System TAE
Plates: Plates:
Silica gel G, 250 μm m thick.
Mobile phase: phase:
Methanol.
Reference compounds: compounds:
Codeine Rf 20,
Trimipramine Rf 36, Hydroxyzine Rf 56,
Diazepam Rf 82.
4/27/2009 ٩٣

System TAF
Plates: Plates:
Silica gel G, 250 μm m thick.
Mobile phase: phase:
Methanol:n-butanol Methanol: butanol (60:40) and
0.1 mol/L NaBr.
Reference compounds: compounds:
Codeine Rf 22,
Diphenhydramine Rf 48, Quinine Rf 65,
Diazepam Rf 85 .
4/27/2009 ٩٤

Location reagents for systems TA, TB
and TC
Ninhydrin spray
Spray the plate with the reagent and then heat
in an oven at 100° 100 for 5 min. Violet or pink
spots are given by primary amines and yellow
colours
Ninhydrin Spray: add 0.5 g of ninhydrin to
10 mL of hydrochloric acid and dilute to
100 mL with acetone. Prepare daily.
4/27/2009 ٩٥

FPN reagent
Red or brown-red brown red spots are given by
phenothiazines and blue spots by
dibenzazepines. This reagent may be used to
overspray a plate which has been previously
sprayed with ninhydrin spray.
FPN Reagent: mix together 5 mL of ferric
chloride solution, 45 mL of a 20% w/w
solution of perchloric acid, and 50 mL of a
50% v/v solution of nitric acid.
4/27/2009 ٩٦

Dragendorff spray
Yellow, orange, red-orange, red orange, or brown-orange brown orange spots
are given by tertiary alkaloids. This reagent may be
used to overspray a plate which has been previously
sprayed with ninhydrin spray and FPN spray.
Dragendorff Spray: (a) mix together 2 g of bismuth
subnitrate, subnitrate,
25 mL of acetic acid, and 100 mL of
water; (b) dissolve 40 g of potassium iodide in
100 mL of water. Mix together 10 mL of (a), 10 mL
of (b), 20 mL of acetic acid, and 100 mL of water.
Prepare every 2 days.
4/27/2009 ٩٧

Acidified iodoplatinate solution
Violet, blue-violet, blue violet, grey-violet, grey violet, or brown-violet brown violet spots
on a pink background are given by tertiary amines
and quaternary ammonium compounds. Primary and
secondary amines give dirtier colours. This solution
may be used to overspray a plate which has
previously been sprayed with ninhydrin spray, FPN
reagent and Dragendorff spray.
Iodoplatinate Solution, Acidified: add 5 mL of
hydrochloric acid to 100 mL of iodoplatinate
solution.
4/27/2009 ٩٨

Mandelin’s Mandelin s reagent
This reagent is preferably poured onto the
plate because of the danger of spraying
concentrated acid. Many different colours are
given with a variety of drugs
Mandelin's Reagent: dissolve 0.5 g of
ammonium vanadate in 1.5 mL of water and
dilute to 100 mL with sulfuric acid. Filter the
solution through glass wool.
4/27/2009 ٩٩

Marquis reagent
This reagent is preferably poured onto the
plate because of the danger of spraying
concentrated acid. Black or violet spots are
given by alkaloids related to morphine. Many
different colours are given with a variety of
drugs
Marquis Reagent: mix 1 mL of
formaldehyde solution with 9 mL of sulfuric
acid. Prepare daily.
4/27/2009 ١٠٠

Acidified potassium permanganate
solution
Yellow-brown Yellow brown spots on a violet background
are given by drugs with unsaturated aliphatic
bonds. bonds
Potassium Permanganate Solution,
Acidified: a 1% solution of potassium
permanganate in 0.25 M sulfuric acid.
4/27/2009 ١٠١

Gas Chromatography.
System GA—amfetamine GA amfetamine RI 1125, amfetamine-TFA amfetamine TFA RI 1095,
amfetamine-PFP amfetamine PFP RI 1330, amfetamine-TMS amfetamine TMS RI 1190,
amfetamine-AC amfetamine AC RI 1501, art (formyl) RI 1100, M (3-OH (3 OH-)-
PFP2 RI 1520, M (3-OH (3 OH-)-TMS2 TMS2 RI 1850, M (3-OH (3 OH-)-AC2 AC2
RI 1930, M (4-OH (4 OH-) ) RI 1480, M (4-OH (4 OH-)-AC2 AC2 RI 1900, M
(3,4–di (3,4 di-OH OH-)-AC3 AC3 RI 2150, M (OH-methoxy
(OH methoxy-) ) RI 1465, M
(OH-methoxy
(OH methoxy-)-AC2 AC2 RI 2065, M (desamino–oxo
(desamino oxo-OH OH-)-AC AC
RI 1520, M (desamino–oxo
(desamino oxo-OH OH-methoxy methoxy-) ) RI 1510, M
(desamino–oxo
(desamino oxo-OH OH-methoxy methoxy-)-AC AC RI 1600, M (desamino– (desamino
oxo–di oxo di-OH OH-)-AC2 AC2 RI 1735; system GB—amfetamine GB amfetamine RI
1150; art (formyl) RI 1142; system GC—RI GC RI 1536; system
GF—RI GF RI 1315; system GAK—retention GAK retention time 4.9 min.
4/27/2009 ١٠٢

Column: DB-5 DB 5 fused silica (30 m × 0.25 mm
i.d., 0.25 μm m film thickness). Column
temperature: 70° 70 for 1 min, ramp to 100° 100 at
30°/min, 30 /min, and to 270° 270 at 10°/min. 10 /min. Injector
temperature: 280°. 280 . Carrier gas: helium, flow
rate 0.8 mL/min. MS detection. Retention
time: 6.5 min.
4/27/2009 ١٠٣

High Performance Liquid
Chromatography.
System HA—k HA k 0.9; system HB—k HB k 8.48;
system HC—k HC k 0.98; system HX—RI HX RI 244;
system HAA—retention HAA retention time, 3.7 min; system
HBC—retention HBC retention time 2.1 min; system HBD— HBD
retention time 3.7 min.
4/27/2009 ١٠٤

Column: Chiralcel OD-RH OD RH (150 × 2 mm i.d.,
5 μm) m) at 35°. 35 . Mobile phase: phosphate–citrate
phosphate citrate
buffer (pH 4.0) with sodium
hexafluorophosphate (0.3 M):acetonitrile
(43:57), flow rate 0.1 mL/min. Fluorescence
detection (λex=330 ( ex=330 nm, λem=440 em=440 nm).
Retention time: 24.6 min.
4/27/2009 ١٠٥

Infra–red Infra red Spectrum
Principal peaks at wavenumbers 700, 740, 1495, 1090,
1605, 825 cm−1
cm
4/27/2009 ١٠٦

Definitive De nitive Identification
Identi cation of
Amphetamines
GC–MS GC MS is the preferred method for the identification
identi cation
of amphetamines. The discussion below centres on
the analysis of amphetamine itself, although the same
principles can also be applied to other members of
this class of drug. However, there are a number of
problems associated with the gas chromatographic
analysis of amphetamine. Being highly polar in
nature, this compound is liable to poor
chromatographic behaviour and tailing if the
analytical instrument is not scrupulously clean
4/27/2009 ١٠٧

4/27/2009 ١٠٨

4/27/2009 ١٠٩

Furthermore, the highly polar nature of the
amino group results in sorption of
amphetamine to the surfaces of the GC system
components. This, coupled with the often low
concentration of the amphetamine in the
sample, results in the false impression that
there is no amphetamine present in the
specimen under investigation
4/27/2009 ١١٠

In order to alleviate this problem,
derivatization can be employed. One of the
easiest processes, for the analysis of
amphetamine, is to derivatize directly with
carbon disulfide disul de and it is this method which
finds nds wide application in the United Kingdom.
For bulk and trace samples, this is achieved by
dissolving the material
4/27/2009 ١١١

4/27/2009 ١١٢

The reaction (see equation (2.1)) is a simple,
pre-column pre column derivatization, derivatization,
involving the amino
group of the amphetamine and the CS2
.Thisprocess Thisprocess reduces the polarity of the
product, improving its chromatographic
behaviour and hence the sensitivity of the
method. In addition, it results in a molecule
which produces characteristic fragments from
the ionization process:
4/27/2009 ١١٣

4/27/2009 ١١٤

Quantification
Quanti cation of Amphetamines
Due to the nature of the compounds being
considered and the need for derivatization,
derivatization,
GC–MS GC MS is not considered the best technique
for sample quantification
quanti cation
4/27/2009 ١١٥

There are a number of difficulties dif culties encountered with
quantification
quanti cation after employing derivatization. derivatization.
These
include the fact because derivatization is another
handling stage in the analytical process, there is
always the risk of sample contamination.
Furthermore, the assumption is made that the
derivatization reactions are ‘complete complete’ and that the
corresponding derivatives are stable for the period
between derivative formation and analysis. Further
factors are that dilutions need to be extremely
accurate and precise to obtain reliable numerical data
and that derivatization can potentially lead to
increases in numerical errors for such data .
4/27/2009 ١١٦

The amphetamines (standards and samples) should be
dissolved in methanolic HCl (100 ml of methanol to
which 175 µl l of concentrated HCl has been added). A
range of standard solutions should be prepared in
order to give a range of concentrations above and
below that which the street sample is thought to
contain, remembering that the latter may only contain
between 0 and 5 wt% amphetamine. If necessary, the
materials (particularly the case samples) should be
sonicated and, following this, centrifuged to remove
any solid materials. The supernatant is retained for
subsequent analysis.
4/27/2009 ١١٧

Having collected the data, a calibration curve should be
plotted. Since amphetamine is frequently synthesized in dirty
apparatus in ‘clandestine clandestine’ laboratories, it may not be possible
to determine which salt form of the drug is present. The
standard is generally supplied as the sulfate form, of the
general formula (amphetamine sulfate). This means that for
every gram of amphetamine sulfate, 73% will be present as the
amphetamine free base. The calibration curve should be
plotted as (UV detector) response against concentration of
amphetamine free base
4/27/2009 ١١٨

Derivatization
4/27/2009 ١١٩

Derivatization results in good chromatographic
behavior and provides compounds for which
distinctive mass spectra can be obtained
This method also has the advantage that the most
common adulterant of amphetamine, namely caffeine,
can also be identified identi ed in this system, and a suitable
mass spectrum obtained As with all drug
identification
identi cation approaches, if the retention time data
and mass spectra of the compounds match, then an
identification
identi cation can be inferred.
4/27/2009 ١٢٠

Mass Spectra
44 91 40 42 65 45 39 43 Amfetamine
44 122 78 121 65 107 91 134
Methoxyamfetamine
44 136 51 135 77 42 78 45
Methylenedioxyamfetamine
44 138 122 137 121 91 78 45
Methylthioamfetamine
4/27/2009 ١٢١