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Qualitative analysis of narcotic compounds from urine

samples of drug addicts using TLC and FTIR

Article  in  International Journal of Medical Toxicology and Legal Medicine · June 2014

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 International Journal of Medical Toxicology & Legal Medicine Vol. 14 No. 1, July -Sep 2011
RESEARCH ARTICLE
ARTICLE
QUALITATIVE ANALYSIS OF NARCOTIC COMPOUNDS
FROM URINE SAMPLES OF DRUG ADDICTS USING TLC
AND FTIR
Anamdeep1, Sandeep Singh Sahota2, Rakesh Kumar Garg3
Abstract
The use of narcotics related compounds (Opium, morphine,
codeine, Heroin etc) is now increasing particularly in
Punjab due to drug trafficking from Golden Crescent
(Pakistan, Afghanistan and Iran). In the present study
the twenty drug addicts admitted in the drug
rehabilitation centers were analyzed by collecting their
urine samples with passage of time. The urine samples
were extracted by solvent extraction technique (Clarke,
1986) and centrifuged. The supernatant were used to
analyze the sample using Thin layer chromatography
(TLC) and Fourier Transform Infrared spectroscopy (FTIR)
with Triglycine Sulfate as detector to determine the drugs
in the urine sample.. This preliminary study shows that
the narcotics substances can be detected in urine using
TLC and FTIR analysis with lapse of time.
Keywords: Narcotics; Morphine; TLC; FTIR;
Qualitative; Urine
Introduction
Narcotics drugs are that substance that depresses
central nervous system functions, bringing sleep
and lessening pain. Thin layer chromatography
has also been primary detection technique for
1. Research Student, Department of Forensic Science,
Punjabi University, Patiala.
2. Junior Research Fellow-UGC, Department of
Forensic Science, Punjabi University, Patiala.
3. Professor and Head, Department of Forensic
Science, Punjabi University, Patiala.
Corresponding Author:
Dr Rakesh Kumar Garg
Department of Forensic Science, Punjabi University,
Patiala. 147002
Phone: +91-175-304-6271
Email: rkgvpbi@yahoo.co.in
77
opiate alkaloids screening [1, 2, and 3]. Fourier
transform infrared spectroscopy (FTIR) is used
as confirmatory test in this study for the
identification of compounds with Triglycine
Sulfate as detector. Triglycine sulfate (TGS) is a
very sensitive detector type for infrared
spectrometers that take measurements in the mid-
infrared (mid IR) range4.
Various authors has conducted studies on
different biological samples (blood, urine and
saliva) and found some notable differences in
the sensitivity of these techniques5,6. The present
study is a pivot work to detect the drug in the
urine sample using TLC and FTIR to determine
the drugs in the body fluids to the minimal level
and with a passage of time.
Materials and Methods
Drug Standards: Poppy husk, morphine was
obtained from drug addicts for the purpose of
comparison and confirmed by instrumental
techniques.
Sample Collection: Human urine samples were
collected from the addicted persons in the drug
rehabilitation centers and hospitals of Patiala.
Twenty (20) samples of urine were collected from
addicted individuals at regular interval of time.
The urine samples were preserved with sodium
fluoride or sodium azide and refrigerated
immediately.
Extraction: To 5ml of the urine sample in a 10ml
centrifuge tube added 0.5ml of 5M Potassium
hydroxide, 3g of Sodium chloride, and 2ml of
ether containing 10µg/ml of diphenylamine.
Mixed thoroughly for 10 min on a rotary mixer.
Centrifuged, and removed about 1ml of ethereal
 International Journal of Medical Toxicology & Legal Medicine Vol. 14 No. 1, July -Sep 2011

layer, and dried with 0.1g of anhydrous sodium The detailed Rf values for solvent system no.
sulfate (Clarke, 1986). 7 is shown in table no II. There results indicate
that they could be separated using these solvents systems
Thin Layer Chromatography: Thin layer
as they were clear and distinct from each other.
chromatography was performed on the smooth
glass plates of 20X10 cm size with Silica gel-G Table 1
(Emerck) used as the adsorbent. The plates were Showing different solvent systems
activated in the oven at about 110ºC for about applied
1hour. The plates were then removed from the
oven and allowed to cool. The plates were then S. No Solvent System Ratio
sprayed with Potassium Hydroxide solution 1 Methanol: 100:1.5
before spotting. The capillary of the same Ammonia
diameter was used in all the experiments to ensure
that approximately the same amount of a 2 Ethyl Acetate: 85:10:5
particular compound was spotted on the plates. Methanol
The spots were made at an equal distance (about 3 Chloroform: 90:10
2 cm) from the bottom & the distance between Methanol
seven parallel spots was also kept constant. The
spots were allowed to dry before putting them to 4 Methanol: 95:5
the particular solvent systems. Ammonia

The plates were developed in an unsaturated 5 Ethyl Acetate: n- 60:35:5

chamber with cyclohexane:chloroform: Butanol: Ammonia
Diethylamine (50:40:10) and air dried and later 6 Chloroform: 60:30:10
exposed to Iodine fumes for visualization after Methanol:
this the plates were sprayed with dragondroff ’s Diphenylamine
reagent.
7 Cyclohexane: 50:40:10
Fourier Transform Infrared Spectroscopy (Ftir): Chloroform:
Sample was grounded with pestle and mortar and Diethylamine
mixed with powdered spectroscopic grade KBr
pellet. After making the pellet it was placed in 8 Diethyl ether: 5:50:40:5
the pellet holder and was scanned sixteen times Acetone: Ethanol
to get fine peaks and spectra was obtained. The
9 Toluene: Acetone: 45:45:7:3
detector used was Triglycine Sulfate (TGS).
Ethanol: Ammonia
Results and Discussions 10 Ethyl Acetate: 70:15:10:5
Cyclohexane:
In the present study, twenty (20) urine samples
Methanol: Ammonia
were examined. Various solvents were tried as
shown in Table 1. In case of analysis of samples under Fourier
The solvents system S#7 (Cyclohexane: Transform Infrared Spectroscopy (FTIR), all the
Chloroform: Diethylamine) and solvents system urine samples showed the presence of strong IR
S#10 (Ethyl Acetate: Cyclohexane: Methanol: band in the region 3818cm-1, 3649cm -1 and
Ammonia) were found to be the most suitable 3410cm-1 which clearly indicates the presence
for the separation of various narcotic compounds of OH functional group and shows
present in the urine. The hRf value of standard intermolecular H-bond. Besides that it also
sample of opium were different (16, 35, 8) poppy showed moderate IR adsorption band at 1622cm-
1
husk (18, 75). and 1621cm -1 , showing the presence of
aromatic stretching present in the morphine,

78
 International Journal of Medical Toxicology & Legal Medicine Vol. 14 No. 1, July -Sep 2011

Table 2
Showing hRf value of samples using
solvent system 7 (Cyclohexane:
Chloroform: Diethylamine)

Sample No. Observations

No. of Spots hRf
R1 4 67, 75, 8, 95
R2 2 79, 92
R3 1 9
R4 1 9
R5 1 87 Figure 1: Showing absorption spectra of urine
sample analyzed using FTIR
R6 2 85
R7 2 82, 95 detected from urine samples in drug addict
individuals with lapse of time. Solvent System
R8 2 79 (Cyclohexane: Chloroform: Diethylamine) is
R9 1 78 more suitable for the separation of narcotic
R10 1 95 compounds and detection of functional group
through FTIR analysis, it can reveal the structure
R11 1 8
of the compounds. The study shows that a
R12 2 35, 8 preliminary screening of the narcotics from urine
R13 1 7.5 samples of drug addicts can help in detection of
these after a lapse of 15 days using TLC and
R14 2 2, 82
FTIR analysis and could be used in investigations.
R15 0 0 It is expected that the study will provide useful
R16 1 8 information to the forensic toxicologist working
in the field that TLC and FTIR techniques can
R17 0 0
provide preliminary information for their
R18 1 9 presence. Further work using different extraction
R19 1 9 procedures and instrumental techniques is in
progress.
R20 1 85
S1 3 16, 35, 8 Acknowledgements
S2 2 18, 75 The authors acknowledge Saket Hospital, Patiala
and Rajindra Government Hosiptal, Patiala for
codeine, thebaine etc. The adsorption band at their permission to collect urine samples of drug
1086cm-1, 1070cm-1 shows the presence of ether addicts. We are also thankful to the each and
linkage in thebaine and codeine. The moderate every individual who provided the samples for
adsorption band at 812cm-1, 809cm-1, 804cm-1 the present study.
and 800cm -1, showed the presence of ortho
substitution in aromatic ring of the compound References
(Fig 1).
1. Clark EGC. (1986) Clarkes’ Isolation and
It is concluded from the present study on identification of drugs. 2nd Edition, Pharmaceutical
narcotic related compound that these can be Society of Great Britain, UK.

79
 International Journal of Medical Toxicology & Legal Medicine Vol. 14 No. 1, July -Sep 2011
2. F Tames, I. D. Watson, W. Morden and I. D. Wilson
Detection and identification of morphine in urine
extracts using thin-layer chromatography and tandem
mass spectrometry. Journal of Chromatography B,
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3. J. P. Comer and Comer L. Applications of thin layer
chromatography in Pharmaceutical analyses. Journal
of pharmaceutical sciences, 1967;56:413-416.
4. M Banan, RB Lal, A Batra. Modified triglycine
sulphate (TGS) single crystals for pyroelectric
infrared detector applications, Journal of materials
sciences, 1992;27: 2291-2297.
80
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5. M Josefsson, R Kronstrand, J Andersson, M Roman.
Evaluation of electrospray ionisation liquid
chromatography–tandem mass spectrometry for
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