SCENARIO 3 BLOCK 4

S, a pharmacist who works at the Police Forensic and Toxicology Laboratory received a tablet sample
from a raid location at a night club in Sleman Regency. It is suspected that the tablet is a psychotropic
type of Amfetamine or its derivatives. To complement the police report, qualitative and quantitative
analyzes were carried out on the tablets.

STEP 1: CLARIFICATION OF TERMS

1. Amphetamines: a stimulant drug that is usually used only to treat hyperactive disorders due to
inattention or attention deficit disorder in adults and children
2. Toxicology: the study of the work and the harmful effects of chemicals or toxins on the
biological mechanisms of an organism
3. Psychotropic: a natural or synthetic substance or drug that is not narcotic and has
psychoactive properties through a selective effect on the central nervous system so that it can
cause characteristic mental activity and behavior
4. forensic lab: a testing laboratory that assists in uncovering a criminal case

STEP 2: PROBLEM FORMULATION

1. Describe the structure of amphetamines and their derivatives?

2. How do amphetamines work?
3. How to analyze amphetamines using the LC-MS / MS instrument?
4. Describe the type of psychotropic drugs?
5. What instruments do you use to analyze amphetamines?
6. What tests are carried out on drugs in the forensic laboratory?
7. What are the side-effects of amphetamines?
8. How are amphetamines actually used in the health sector?
9. What are the medical benefits of psychotropics?
10.How is the relationship between psychotropics and amphetamines?
11.What types of psychotropics other than amphetamines?

STEP 3: BRAINSTROMING

1. Amphetamines have the chemical name 1 phenyl 2 amino propane or phenyl isopropyl.
Amphetamine dissolves in water (1:50) is a solution that is colorless and alkaline to litmus,
tastes to burn the tongue, at temperature it slowly evaporates, dissolves well in ethanol, ether
and chloroform melting point 200-203 degrees Celsius.
2. The mechanism of action of amphetamines on the nervous system is influenced by the release
of biogenic amines, namely dopamine, norepinephrine, or serotonin or the release of all three
from storage sites in the preparation which are located at nerve endings (Japardi, 2002).
3. The amphetamine analysis method was performed using the LC-MS / MS instrument. The LC
instrument consists of the LC-20AD XR UFLC system with a SIL-HT automatic sample
injector (Shimadzu, Kyoto Japan). The LC-MS / MS system is controlled with the Analyst
software, version 1.6.3 (Applied Biosystems).
4. Group I: Psychotropics that can only be used for scientific purposes and are not used in
therapy, and have a strong potential to cause addiction syndromes. Example: Ecstasy.

Group II: Psychotropics that have medicinal properties and can be used in therapy and / or for
scientific purposes and have a strong potential to cause addiction syndromes. Example:
Amphetamine.

Group III: Psychotropics that have medicinal properties and are widely used in therapy and /
or for scientific purposes and have moderate potential to cause addiction syndromes.
Example: Phenobarbital.

Group IV: Psychotropics that have medicinal properties and are very widely used in therapy
and / or for scientific purposes and have a mild potential to cause addiction syndromes.
Example: Diazepam, Nitrazepam (BK, DUM).
5. Amphetamine analysis was performed using the LC-MS / MS instrument. The LC instrument
consists of the LC-20AD XR UFLC system with a SIL-HT (Shimadzu,
Kyoto Japan)
6. Forensic laboratories are often called in to identify powders, liquids, and pills that may be
illegal drugs. Basically, there are two categories of forensic tests which are used to analyze
drugs and other unknown substances, namely:
- The presumptive test
- Color test
- Ultraviolet spectrophotometric test
 - Microcrystalline test
7. The side effect of using amphetamines is the appearance of feeling very tired after the effects
wear off within a few hours. Long-term use causes dependence and intolerance so that users
will always want to take the drug to prevent withdrawal effects (withdrawal). In the case of
using excessive doses, it will cause life-threatening conditions (Substance Abuse and Mental
Health Services Administration, 2013a, 2013b).
8. Amphetamines are drugs used to treat narcolepsy disorders, attention deficit disorder with
hyperactivity (ADHD), Parkinson's disease, and obesity.
9. In the medical field, several types of psychotropic drugs are used for the treatment of certain
mental disorders, such as depression, anxiety disorders, bipolar disorder, sleep disorders, and
schizophrenia.
10.Amphetamine is a synthetic compound analogous to epinephrine and is an indirect
ketecholamine agent (Japardi, 2002). Amphetamines are included in group II psychotropic
substances (Hawari, 2006).
11.* Sedatin.
* Rohypnol.
* Valium.
* Metakualon.
* Phenobarbital.
* Shabu-shabu.
* Ecstasy.
* Ritalin
* Methylphenide

STEP 4: PROBLEM ANALYSIS

Amphetamine
Identification
 The use or dosage of
Amphetamines
amphetamines and
drugs according to the
rules and laws

Molecular formula
Use in the medical field

Group and relationship

with psychotropics
Quantitative and
qualitative analysis

Amphetamine
derivatives and
characteristics Testing conducted on
drugs

Amphetamine
mechanism of action

Amphetamine side
effects

Indications and benefits

of amphetamines

1. -3.4 methylenedioxyamphetamine (MDA)

-3.4 methylenedioxymetamphetamine
-2.5 dimethoxy -4 ethylamphetamine
-4 bromo 2,5 dimethoxyamphetamine
 -3,4,5 trimethoxyamphetamine (TMA)
-2.5 dimethoxy 4 methylamphetamine (DOM)

2. In dopamine it was found that amphetamine inhibits dopaminergic and synapstosome re-
uptake in the hypothalamus and directly releases newly synthesized dopamine (Japardi, 2002).

In norepinephrine, amphetamine blocks norepinephrine re-uptake and also causes the release
of new norepinephrine, the addition or reduction of carbon between the phenyl and nitrogen
rings attenuates the effect of amphetamine on the release of norepinephrine re-uptake
(Japardi, 2002).

Whereas in serotonin, methamafetamine deviate with strong electrons that dance to replace
the phenyl ring will affect the serotoninergic system (Japardi, 2002).

These three biogenic receptors work mutually influence one another. The activity of the
central nervous system that occurs through these pathways in the brain, each of which gives
rise to the activity and personality of the individual user. Stimulation of the motor center in
the forebrain media area (medial forebrain) causes an increase in norepinephrine levels in the
synapse, causing euphoria and increasing libido (Japardi, 2002).

Stimulation of the ascending reticular activating system causes an increase in motor activity
and reduces fatigue (Japardi, 2002). Stimulation of the dopaminergic system in the brain
causes symptoms similar to schizophrenia (Japardi, 2002).

The conclusion is the work of the three receptors mentioned above, can cause euphoria,
increase libido, increase motor activity, reduce fatigue and cause symptoms similar to
schizophrenia for amphetamine users.

3. For the spectrometer we use the Linear Ion Trap Quadrupole LC-MS / MS. The procedure for
taking a sample is that 5 mL of human blood is put into an EDTA tube. The blood cells were
separated from the plasma by means of a centrifugation procedure for 10 minutes at a rate of
2,000 x g. After centrifugation, the plasma was transferred to a sterile polypropylene tube
using a Pasteur pipette. Samples were stored in an ice cupboard at -20 ° C until the time of
analysis. Sample preparation was carried out by using a protein precipitation extraction
technique. The frozen plasma samples were thawed at room temperature. After the liquid
plasma sample, the plasma is cortexed to ensure that all the ingredients are completely mixed.
Furthermore, 100 µL of plasma was added with 300 µL of acetonitrile. The mixture was
cortexed for 5 seconds and centrifuged for 2 minutes at a speed of 14800 rpm. The
supernatant was filtered with a 0.2 µm diameter filter, then transferred to a new vial. A total
of 8 microliter samples were injected into LC-MS / MS for detection of amphetamine content.
 This study succeeded in finding an effective, efficient, and simple procedure (protein
precipitation) that can detect amphetamines in human blood plasma (Liquid Chromatography
Tandem Mass Spectrometry / LC-MS / MS). This method can get results in a short period of
time, at a low price and a small sample size. This method also has excellent sensitivity,
specificity, and recovery for detection of amphetamines in human blood plasma, at
concentrations in excess of 100 ng / mL. This method is recommended for confirmatory
testing, which is needed to confirm the presence of amphetamines in human blood plasma

4. Examples of class 1 psychotropics include LSD, DOM, Ecstasy, and others. These drugs will
have a hallucinatory effect on the user and change feelings drastically.

Examples are Methamphetamine and Phenytoin

Examples of group 3 substances include Mogadon, Buprenorphine, Amobarbital,

Some types include Lexotan, Koplo pills, sedatives or sedatives, hypnotics or sleeping pills,
Diazepam, Nitrazepam

5. The LC-MS / MS system is controlled with the Analyst software, version 1.6.3 (Applied
Biosystems). The analytical columns used were Agilent, Eclipse Plus Phenyl-Hexyl (150 mm
x 2.1 mm, 5µm). The column temperature was 40 ° C for a total analysis time of 10 minutes.
The mobile phase used was 10 mM ammonium formate in water (pH 6.6) at pump A and
0.1% FA (in acetonitrile) at B. Flow rate was 0.3 mL / minute with an elution gradient set
according to room temperature. The gradient starts with 5% B, then increases to 40% B at 3
minutes, and lasts up to 4 minutes. Subsequently the gradient rose to 100% B at 6 minutes,
and lasted up to 8 minutes. The gradient then returned to 5% B at 8.01 minutes and this
condition lasted for 10 minutes. The sample volume at injection was 8 µL. For the
spectrometer we use the Linear Ion Trap Quadrupole LC-MS / MS Spectrometer, QTRAP
5500, with ESI probe. This instrument is operated with a negative ionization model. Some of
the optimal spectometric parameters used for amphetamine detection are the ion source
temperature of 4500C, the ion spray voltage of 5500 V, the curtain gas pressure of 20 psi, the
1 ion source gas pressure of 35 psi, and the ion source gas pressure of 2 of 35.0 psi.

6. Presumptive tests, such as color tests, only give an indication of what type of substance is
present. However, forensic experts were unable to specifically identify the substance. With
confirmatory tests, such as gas chromatography / mass spectrometry can get more specific
results and determine the exact identity of a substance.

Color test: This test can expose an unknown drug to a chemical or chemical mixture. The
color change of the substance can help determine the type of medication that is present. For
 example, on a cobalt thiocyanate test, if the substance turns the liquid blue, it means the drug
is heroin.

Ultraviolet spectrophotometric test, which analyzes how substances react to ultraviolet (UV)
and infrared (IR) light. Spectrophotometric machines emit UV and IR light, and then measure
how the sample reflects or absorbs these rays to provide an overview of what types of
substances are present.

Microcrystalline test: This method is done by adding a drop of the suspect material to the
chemical on a microscope slide. The mixture will start to form crystals. Each drug has its own
crystal pattern when viewed under a polarized light microscope

7.-Effects of acute use of amphetamines

The effects that can occur after using amphetamines depend on the amount of amphetamine
consumed and the way it is administered. In general, the use of amphetamines has an acute
effect in the form of disorders of the autonomic sympathetic nervous system, such as
hypertension, tachycardia, hyperthermia, tachypnea, and vasoconstriction. In addition, acute
use of amphetamines can cause euphoria, increased energy and alertness, increased libido and
self-confidence, a feeling of increased physical and mental capacity, and increased
productivity.

-Effects of amphetamines on cognitive function

The effect of amphetamine on brain function is related to the release of dopamine,

norepinephrine, and serotonin. These three neurotransmitters are produced in neuron cells
located in the midbrain and brainstem and are projected to almost all parts of the brain.
Amphetamine users

Various studies have shown an increase in cognitive performance, particularly information

processing speed, psychomotor function and attention, with acute administration of
amphetamines in therapeutic doses, but still a high risk of experiencing dependence.

Amphetamine users can also overdose. Overdose is an emergency situation which, if not
handled properly, can lead to heart attack, stroke, rhabdomyolysis, kidney failure, and even
death.

8. Amphetamines are currently used by the medical community to treat several conditions,
including narcolepsy and ADHD. The State University of New York reports that in some
cases, they have also been shown to be effective in treating depression and obesity. One of the
most uses
 surprising to amphetamines is the use of the drug helps stroke victims to recover more
quickly.

A recent study by the Karolinska Institute of Sweden showed that treatments can be of great
help for those who have had a weak stroke

9. Medical Benefits of Psychotropics

Medically and legally, psychotropic drugs should only be used according to a prescription and
supervision from a specialist. These drugs are usually used to treat various conditions or
certain diseases, such as:

• Mental or psychological disorders

• Seizures or epilepsy

• Parkinson's disease

• Sleep disturbances, such as insomnia or narcolepsy

• Chronic fatigue syndrome

In addition, psychotropic drugs are also often used as anesthetics or anesthetics to prevent and
treat severe pain due to certain medical actions, such as surgery. However, unfortunately,
these drugs can also be abused. If not used as indicated, drugs or psychotropic substances can
cause dangerous addictive effects and even death.

Because of its addictive effects (addiction), psychotropic drugs should only be used for
medical purposes based on a doctor's prescription.

10. Psychotropic is a substance or drug, both natural and synthetic, not narcotics, which has
psychoactive properties through selective influence on the central nervous system which
causes distinctive changes in mental activity and behavior (Japardi, 2002).

Class II psychotropics are psychotropic substances that can only be used for scientific
purposes and are not used in therapy and have a very strong potential to cause dependency
syndrome (Ministry of Health, 2010).

Amphetamines are a group of stimulants (Ministry of Health, 2010). The stimulant class is a
type of drug that can stimulate bodily functions and increase work excitement. Amphetamines
are divided into two types, namely MDMA (Methylene dioxy methamphetamine) and
amphetamines. Amphetamine has a longer working duration than MDMA, and has a stronger
hallucinatory effect (Ministry of Health, 2010).
 Shabu or amphetamine is a group of narcotics which is a nervous system stimulant with the
current name methamphetamine hydrochloride, which is a derivative of the nerve stimulant
amphetamine (Japardi, 2002). Shabu is in the form of white crystals similar to MSG (Mitra
Bintibmas, 2010).

11. 1. Stimulant drugs

This type of psychotropic is a stimulant drug that can stimulate nerves so that it can have a
more confident effect. Many types of psychotropic substances including stimulant drugs,

for example: caffeine, cocaine, marijuana, and amphetamines. Amphetamines are usually
found in ecstasy pills.

2. Depressant Drugs

This type of psychotropic is a depressant drug that can have an effect, namely reduced
nervous system work, decreased awareness, and drowsiness. Types of substances that include
depressant drugs,

examples: alcohol, sedatin or BK pills, Magadon, Valium, and Mandrak (MX), Cannabis and
Barbiturates.

3. Hallucinogenic drugs

This type of psychotropic is a hallucinogenic drug that can cause hallucinations, namely
hearing or seeing something that is not real.

For example: namely Licercik Acid Dhietilamide (LSD), psylocibine, micraline and
marijuana.

STEP 5: LO

1. What are the categories of forensic tests used to analyze unknown drugs or other substances?
2. Which analytical method is used to analyze amphetamines?
3. Why can this compound be analyzed by gas chromatography?
4. How to instrument the gas chromatography method?
5. What are the mobile and stationary phases used in gas chromatography?
6. Advantages and disadvantages of using gas chromatography?
7. What detectors are used in gas chromatography and how do I read the output?
8. Any efforts that can be made to increase the effectiveness of the analysis?

STEP 6: LEARN INDEPENDENT

STEP 7: REPORTING OF INDEPENDENT LEARNING RESULTS

1. Drug Forensic Testing
Forensic laboratories are often called in to identify powders, liquids, and pills that may be
illegal drugs. Basically, there are two categories of forensic tests which are used to analyze
drugs and other unknown substances, namely:
Presumptive tests, such as color tests, only give an indication of what type of substance is
present. However, forensic experts were unable to specifically identify the substance. With
confirmatory tests, such as gas chromatography / mass spectrometry can get more specific
results and determine the exact identity of the substance.

Color test: This test can expose an unknown drug to a chemical or chemical mixture. The
color change of the substance can help determine the type of medication that is present. For
example, on a cobalt thiocyanate test, if the substance turns the liquid blue, it means the drug
is heroin.

Ultraviolet spectrophotometric test, which analyzes how substances react to ultraviolet (UV)
and infrared (IR) light. Spectrophotometric machines emit UV and IR light, and then measure
how the sample reflects or absorbs these rays to provide an overview of what types of
substances are present.

Microcrystalline test: This method is done by adding a drop of the suspect material to the
chemical on a microscope slide. The mixture will start to form crystals. Each drug has its own
crystal pattern when viewed under a polarized light microscope.

1. Sample preparation
Several things that need to be taken into account in the sample preparation stage are: the type
and biological characteristics of the specimen, the physicochemical of the specimen, and the
purpose of the analysis. Thus you will be able to design or select a sample handling method,
the number of samples to be used, and choose the right method of analysis. The handling of
the sample deserves special attention, as the majority of the sample is biological material,
thereby preventing the breakdown of the analyte as much as possible.

2. Screening Test
Screening test to filter and identify the class of compounds (analytes) in the sample. Right
here
analytes are classified based on both their physicochemical properties, chemical properties
and pharmacological effects. Narcotics and psychotropic drugs in general in the screening test
are grouped into opiates, cocaine, cannabinoids, amphetamine derivatives, benzodiazepine
 derivatives, tri-cyclic anti-pressure compounds, barbituric acid derivatives, methadone
derivatives. This grouping is based on the structure of the molecular nucleus. For example,
here is taken a compound of the opiate group, where this compound has the basic structure of
morphine, several compounds that have the basic structure of morphine such as heroin, mono-
acetyl morphine, morphine,
morphine-3-glucuronide, morphine-6-glucuronide, acetylcodeine, codeine, codeine-6-
glucuronide, dihydrocodeine and its metabolites, as well as other opiate derivatives that have
a morphine core.
3. Verification Test
This test aims to confirm the identity of the analyte and determine the level. The confirmatory
test is at least as sensitive as the screening test, but should be more specific. Generally, the
verification test uses chromatography techniques in combination with other detector
techniques, such as: gas chromatography - mass spectrophotometry (GC-MS), high
performance liquid chromatography (HPLC) with diode-array detectors, liquid
chromatography - mass spectrophotometry (LC-MS), TLC-Spectrophotodensitometry, and
other techniques. Increasing the level of specificity in this test will make it possible to identify
the identity of the analyte, so that it can determine the specific toxicity that is present.
4. Interpretation of the Analysis Findings
The findings of the analysis themselves have no meaning if the meaning of the findings is not
explained. A forensic toxicologist is obliged to translate these findings based on their
expertise into a sentence or report, which can explain or be able to answer questions that arise
regarding the problem / case that is alleged.
2. Analyzing amphetaine and its derivatives using gas chromatography analysis method. From
the chromatography method, it can be used for qualitative analysis to determine the sample
that is suspected to contain amphetamine by using the comparison of the results of the
standard solution chromatogram and the sample chromatogram through the retention time that
appears.

Another method used in drug examination (amphetamines and derivatives) is Competitive

Immunochromatography, by dipping the strip vertically on the urine specimen then waiting
for a few minutes and seeing the results, if the line is shown on the control and the test shows
negative, if the line is shown on the control it shows positive whereas if it is not listed the line
indicates invalid. So that the results obtained that the urine sample tested showed a positive
result, meaning that the patient is a drug user.

# Place and Time of Implementation

The analysis is carried out at the forensic laboratory
 #Tools and materials
Tool
1. Strip test
2. Dropper
3. Tissue
4. Tube
5. Timer
material
Sample urine

#Work procedures
1. Take a urine sample to be examined.
2. Inserted into the tube to taste.
3. Opened the test strip tool that had been provided.
4. Placed on a flat table.
5. Written sample label.
6. Dip the strip vertically in the urine specimen for 10-15 seconds.
7. Wait for the C and T lines to form on the test strip tool.
8. Read the striptest tool, if only a pink band is formed on Control (C) then the result is
positive,
formed two pink bands on Control (C) and in Test (T) stated negative results, and
invalid tool if no pink band is formed on Control (C) and on Test (T) or
a pink band was formed in Test (T) while in Control (C) there was no band formed.

3. Because it is used in chemical analysis for the separation and analysis of compounds which
can vaporize without decomposition. Common uses of GC include testing the purity of a
particular compound, or separating different components in a mixture (relative levels of the
component can also be determined). In some circumstances, KG can help identify
compounds. In preparative chromatography, GC can be used to prepare pure compounds from
a mixture.

4. GC MS Method
The analytical method using GC MS (Gas Chromatography ¬-Mass Spectroscopy) can
measure the type and content of compounds in a sample both qualitatively and quantitatively.
This instrument is a combination of two instruments, namely Gas Chromatography which
 functions to separate compounds into single compounds and Mass Spectroscopy which
functions to detect types of compounds based on their fragmentation patterns. Measurements
using GC MS are generally limited to compounds in the form of gases or liquids that have a
vapor pressure of at least 10-10 torr.

GC MS Instruments Working Principles

The sample injected into Gas Chromatography will be converted into the vapor phase and
flowed through the capillary column with the help of a carrier gas. The separation of mixed
compounds into single compounds occurs based on differences in chemical properties and the
time required is specific for each compound. The detection takes place in Mass Spectroscopy
with the mechanism of bombarding compounds by electrons into ionized molecules and
recording the fragmentation patterns formed compared to the fragmentation patterns of
standard compounds indicated by the percentage Similarity Index (SI).
5. The mobile phase in GC is also called the carrier gas because its initial purpose is to carry the
solute to the column, hence the carrier gas has no effect on selectivity. The conditions for the
carrier gas are: non-reactive; pure / dry because otherwise it will affect the detector; and can
be stored in high pressure tanks (usually red for hydrogen, and gray for nitrogen.
The stationary phase is a layer of microscopic liquid or polymer on top of the solid support
for the stationary phase, which is in a glass or metal tube called a column.
6. Advantages of Gas Chromatography
a. Short analysis time and high separation sharpness
b. Can use longer columns for high separation efficiency
c. Gas has a low viscosity
d. The equilibrium of the partition between gas and liquid takes place quickly so that the
analysis is relatively fast and the sensitivity is high
e. The use of the liquid phase allows us to choose from a very wide variety of stationary
phases that separate nearly all mixtures.
Disadvantages of Gas Chromatography
a. Gas chromatography techniques are limited to volatile substances
b. Gas chromatographs are not convenient for separating large quantities of mixtures.
Separation at the (mg) level is easy, gram-level splitting is possible, but pound or ton
separation is difficult unless other methods are available.
c. The gas phase compared to most of the liquid phase is not reactive to the stationary phase
and solutes.
7. There are several types of detectors for gas chromatography, including:
a. Flame Ionization Detector (FID), detects almost all organic components
 b. Flame Photometric Detector (FPD), detects components that contain phosphorus and
sulfur
c. Flame Thermionic Detector (FTD), detects organic components that contain phosphorus
or nitrogen
d. Thermal Conductivity Detector (TCD), detects almost all components except the carrier
gas
e. Electron Capture Detector (ECD), detects electrophilic components
f. Mass Spectrometer (MS),

How to read gas chromatography output:

From the GC / MS results, chromatographic retention time data will be obtained with several
compound peaks (the largest multiple can be seen from the highest graph) from the
spectogram data, the fragmentation of each compound is obtained, based on the typical
fragmentation pattern and base peak. each compound can be known.

From the retention time obtained. It is matched with the literature retention time in order to
obtain compound data from chromatography. You can get the% area data which will be used
to calculate the concentration of the substance, after passing it on to gas chromatography it is
passed on to mass spectroscopy to determine the gragmentation of the sample. The mass
spectrum of each compound will be matched with the mass spectrum of the data library

8. One way to increase the effectiveness of the analysis is to perform maintenance on the
instruments used, namely Gas Chromatography Instrument Maintenance.
The treatment steps are as follows:
First, if you are a new user who has never used this tool, be sure to find out the procedure for
using it. You can ask an expert user, read the manual book, or ask for brief training on how to
use it.
Second, provide a UPS in case the power goes out while the machine is running.
Third, be sure to always remember to open and close the carrier gas lever before and after use.
Fourth, do the calibration periodically, especially when the gas chromatography tool
encounters unsuitable peaks.
After completion of use, be sure to turn off the instrument according to procedures, such as
lowering the inlet and column temperatures first.
If you feel that there are (technical) irregularities while the machine is running, be sure to
contact the technician team.