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A Comparison of Red Pigments in Different Lipsticks Using Thin Layer
Chromatography (TLC)
Article in Journal of Analytical & Bioanalytical Techniques · January 2013
DOI: 10.4172/2155-9872.1000157
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Analytical & Bioanalytical
Joshi et al., J Anal Bioanal Techniques 2013, 4:1
http://dx.doi.org/10.4172/2155-9872.1000157
Techniques
Research Article
Open Access
A Comparison of Red Pigments in Different Lipsticks Using Thin Layer
Chromatography (TLC)
Bhawana Joshi1, Kapil Verma1* and Jyoti Singh2
1
2
Forensic Science, Amity Institute of Forensic Sciences (AIFS), Amity University, India
Amity Institute of Forensic Sciences (AIFS), Amity University, India
Abstract
The main aim of present work is chromatographic analysis of red pigment in different well known and local
brands of lipsticks. Lipstick samples of different brands of similar color were selected for this study. Coloring agent
was analyzed by thin layer chromatography (TLC). Using different solvent systems [Toluene/Benzene] (4:12),
Toluene/Benzene/Cyclohexane (4:12:4), Toluene/Benzene/Diethyl ether (4:4). It is hypothesized that through thin
layer chromatography analysis of the red pigment in these different brands will provide no characteristic data to
distinguish among lipstick sources. There is no significant difference in the hRf values among the local and well
known brands of lipsticks which can be used as unique feature.
Keywords: Chromatographic analysis; Red pigments; Lipsticks;
Lipstick samples; TLC; Solvent system; hRf value
Introduction
In forensic science, comparative examinations are usually based
on the physical or chemical nature of a substance, or both. Lipsticks
are composed of waxes, oils, organic dyes [1], and inorganic pigments
[2,3]. Color matching can do identifying the lipstick responsible for
leaving a smear. This color analysis may be used to identify the lipstick
found at the crime scene [4].
The colors of lipstick are often due to a mixture of several pigment
compounds. These pigments can be separated using thin layer
chromatography. Depending on the type of pigment, the mobile phase
will vary. Lipsticks are soluble in toluene, so toluene serves as the mobile
phase. After separation, the chromatogram is complete and illustrates
the different pigments that make up a particular color of lipstick [2,3,510].
Traces of Lipsticks, cosmetics, nail polish, or other smears could be
found left on drinking cups, glasses, cigarette butts, and tissue papers
and may all be significant forensic evidence [11] in the investigation of
a crime, especially in cases such as a sexual assault or a homicide [12].
This physical evidence may be found on clothing, parts of the body, a
tissue, or cigarette [5,13]. By comparing the composition of a lipstick
smear with that of a victim, forensic scientists can demonstrate indirect
proof of contact or a relationship between victim and suspect. Also, it is
sometimes possible to extract saliva DNA from the print and may link
a suspect to a crime scene [2].
Various methods of forensic lipstick analysis were reported [6,8,1417]. Small amount of lipstick (approximately 10 µg) could lead to good
comparisons in TLC [3]. Thin Layer Chromatography (TLC) is a widelyused chromatography technique used to separate chemical compounds.
Thin-layer chromatography incorporates a solid stationary phase and
a moving liquid phase to cause a separation of the constituents of a
mixture. Although simple test may be run by simply allowing a solvent
to pick up a piece of porous paper, a more revealing test requires the
preparation of a plate. Because most compounds are colorless, no
separation will be noticed after development unless the materials are
visualized. This is may be done by:
• Exposing to UV light
J Anal Bioanal Techniques
ISSN:2155-9872 JABT, an open access journal
• Exposing to fluorescent dyes
• Exposing to iodine
• Spraying with a reagent
These procedures may be used alone or in conjunction to make the
components of a sample visible. The distance a component has traveled
up a plate can be assigned a numerical value known as the Rf value.
Rf is defined as the distance traveled by the component divided by the
distance traveled by the solvent. In the example to the right, the solvent
was allowed to travel 10 cm up the plate before the plate was removed
from the chamber and dried.
Materials and Methods
Jar with lid, lipstick samples (Branded and Local), TLC plates,
mobile phase/solvent phase, clean cotton piece, sterilized scissor, fine
capillaries, ruler, pencil, todine pellets, titer plates, graduated cylinder,
gloves, protective mask and thirty samples of lip impressions bearing
lipstick were obtained on cloth using various brands and types of
lipsticks, which were available from volunteers [5,13,18-20].
Thin-Layer Chromatography (TLC)
A thin-layer plate is prepared by coating a glass plate with a thin
film of a granular material. Commonly, silica gel or aluminum oxide or
cellulose immobilized onto a flat, inert carrier sheet is used, but paper
may suffice in simpler experiments. These serve as the solid stationary
phase. If the sample to be analyzed is a solid, it must first be dissolved
in a suitable solvent. A few micro liters of the solution are then applied
*Corresponding author: Kapil Verma, M.Sc Forensic Science, Amity Institute
of Forensic Sciences (AIFS), B-Block, Lower Ground Floor, Amity University,
Uttar Pradesh, Sec-125, Noida-201303, Uttar Pradesh, India, E-mail:
forensic.kapilalert@gmail.com
Received November 22, 2012; Accepted December 17, 2012;
December 24, 2012
Published
Citation: Joshi B, Verma K, Singh J (2013) A Comparison of Red Pigments in
Different Lipsticks Using Thin Layer Chromatography (TLC). J Anal Bioanal
Techniques 4: 157. doi:10.4172/2155-9872.1000157
Copyright: © 2013 Joshi B, et al. This is an open-access article distributed under
the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and
source are credited.
Volume 4 • Issue 1 • 1000157
Citation: Joshi B, Verma K, Singh J (2013) A Comparison of Red Pigments in Different Lipsticks Using Thin Layer Chromatography (TLC). J Anal
Bioanal Techniques 4: 157. doi:10.4172/2155-9872.1000157
Page 2 of 4
to the lower edge of the plate. A liquid sample may be applied directly
to the solid state in the same manner. The plate or paper is then placed
upright into a closed chamber that contains a selected solvent.
from the spotted pencil line. Some lipsticks have only two or three
components, and some have more. Enter these measurements on the
Data Table.
The solvent will slowly begin to pick up the plate by capillary action.
It is the rising solvent that serves as the moving phase in thin-layer
chromatography. As it moves past the sample spot, the components of
the sample will become distributed between the stationary solid phase
and the moving liquid phase. Those components with the greatest
affinity for the moving phase will travel up the plate at a faster speed
as compared to those that have greater affinity for the stationary phase.
13. Determine the Rf for each lipstick component of all lipstick
samples. Enter those values on the Data Table. To calculate the Rf value,
divide the distance traveled by each lipstick component by the distance
traveled by the solvent.
Thin-Layer Chromatography (TLC) is a method use to separate
components from each other in a mixture. It is normally experimented
on glass, aluminum foil or plastic which is coated by some kind of
absorbent material (ex: silica gel, aluminum oxide). It takes the Rf
values of each color component and compares them; Rf values are
experiments that depend on the polarity of the substance on the paper
chromatography [21];
Samples are run for both the samples of branded and local lipstick.
Use hood if possible. Self life is about one month.
Rf =
Distance traveled by one lipstickcomponent from the spotted pencil line
Distance the solvent moved from thespotted pencil
Steps followed are shown in figures 1-11.
Results
The data collected from the chromatograms were recorded, and
1. A piece of cotton cloth was washed in a detergent solution,
immersed in a hot water, and dried.
2. This cloth was cut into small pieces (2 cm×2 cm) and lipstick
smears were rubbed onto these pieces.
3. The stained areas from the cloth pieces were cut and transferred
into serially marked small bowls.
4. These samples were mixed with an extracting solution and each
bowl was then shaken for about 5-10 minutes to remove the stain from
the cloth piece.
Figure 1: Sample of different lipstick for study.
5. Cloth piece is taken out, and the extract was used for further
examination with TLC.
6. Obtain jar with lid, piece of filter paper and a TLC plate. Handle
TLC plate by the edges only, avoid touching the white silica layer.
7. With a pencil and ruler, gently draw a line across the short side
of the TLC plate about 1.5 cm from the bottom of the plate. At even
intervals label the top of the plate with the letters: 1B, 2B, 3B, 4B, 5B,
6B for six different well known brands of red lipsticks on a single TLC
plate. Similarly 1L, 2L, 3L, 4L, 5L, 6L for six different local brands of
red lipsticks.
Figure 2: Samples rubbed on a clean cotton piece Bowls
contains the extracted sample.
8. Using a capillary, place a dot of each lipstick sample along the
bottom penciled line directly under the corresponding label on the
top of the plate. The dots should be about 0.2 cm in diameter and dark
enough to be clearly visible.
9. Using a pipette dispenses mobile phase/solvent in the ratio
(4:12:4) into the jar. The mobile phase/solvent should be about 0.5 cm
deep.
10. Carefully insert the TLC plate into the jar, sample end down.
The lipstick dots must be above the mobile phase/solvent. Secure the
lid.
Figure 3: Picture showing TLC plate.
11. Allow the mobile phase/solvent to rise to within one cm of the
top of the plate (5-10 minutes). Watch–do not allow the mobile phase
to rise to the very top of the plate. Remove the TLC plate and mark the
solvent front with a pencil.
12. Measure the distance the mobile phase/solvent moved in cm
(the distance from the spotted pencil line to the solvent front end). Also
measure the distance in cm each component of the lipsticks moved
J Anal Bioanal Techniques
ISSN:2155-9872 JABT, an open access journal
Figure 4: Extract preparation in Toluene (Bowls contains the
extracted sample).
Volume 4 • Issue 1 • 1000157
Citation: Joshi B, Verma K, Singh J (2013) A Comparison of Red Pigments in Different Lipsticks Using Thin Layer Chromatography (TLC). J Anal
Bioanal Techniques 4: 157. doi:10.4172/2155-9872.1000157
Page 3 of 4
Figure 5: Extract preparation in Toluene (Bowls contains the extracted
sample).
Figure 10: Chromatogram of local lipsticks (Spots seen for local brand of
lipstick when viewed under Iodine fumes).
Solvent System: Toluene/Benzene/Cyclohexane
Figure 11: Chromatogram of branded lipsticks (Spots seen for Branded
brand of Lipstick when viewed under Iodine Fumes).
Figure 6: TLC plate inside the jar containing developing solvent.
Solvent System:Toluene /Acetone
Column 3
Lipstick No. of Distance Lipstick
Samples Spots
Components
Moved (cm)
1L
2L
3L
Figure 7: Jar containing iodine pellets.
Solvent System:Toluene/Benzene/Diethyl ether
4L
5L
6L
Figure 8: Jar containing iodine pellets.
Column 4
Distance
Mobile Phase
Moved (cm)
hRf Values for
Each Colored
Component
1
0.7
10
7
2
1.2
10
12
3
2.7
10
27
4
3.7
10
37
5
4.7
10
47
6
7.6
10
76
1
1.3
10
13
2
2.7
10
27
3
3.7
10
37
4
7.6
10
76
1
0.3
10
3
2
1.3
10
13
3
2.5
10
25
4
4.2
10
42
5
7.9
10
79
1
0.3
10
3
2
1.3
10
13
3
3.7
10
37
4
7.9
10
79
1
1.3
10
13
2
3.6
10
36
3
7.8
10
78
1
0.3
10
3
2
1.3
10
13
3
2.9
10
29
4
3.6
10
36
5
8
10
80
Table 1: Rf values of the red dyes in the local brands of lipsticks.
the Rf values were calculated using distance solute travelled/distance
solvent travelled (dsu/dsv). All of these values are shown in the table.
The Rf values of the red dyes in the local brands of lipsticks are given in
table 1 and in the well known brands are given in table 2.
For local brands
Figure 9: Chromatogram appeared after interaction with iodine fumes
(Spots seen for local brand of lipstick when viewed under Iodine fumes).
J Anal Bioanal Techniques
ISSN:2155-9872 JABT, an open access journal
Table 1 shows the result obtained from the local brands of
lipsticks. In which sample marked as 1L shows six pigments having
Volume 4 • Issue 1 • 1000157
Citation: Joshi B, Verma K, Singh J (2013) A Comparison of Red Pigments in Different Lipsticks Using Thin Layer Chromatography (TLC). J Anal
Bioanal Techniques 4: 157. doi:10.4172/2155-9872.1000157
Page 4 of 4
Column 3
Lipstick No. of Distance Lipstick
Samples Spots
Components
Moved (cm)
1B
1
0.4
2B
1
0.35
2
1.3
3B
1
0.25
2
1.55
4B
1
0.2
2
1.3
5B
1
0.25
2
1.5
6B
1
0.25
2
1.5
Column 4
Distance
Mobile Phase
Moved (cm)
10
10
10
10
10
10
10
10
10
10
10
Rf Values for Each
Colored Component
4
3.5
13
25
15.5
2
13
2.5
15
2.5
15
Table 2: Rf values of the red dyes in the well known brands of lipsticks.
Sample No.
1L
2L
3L
4L
5L
6L
Name & Number
biros 14
biros 8
matte lip color 293
aver matte 147
iffy super soft 15
iffy matte soft 17
Brand
local
local
local
local
local
local
Color
pink
rosy red
red
light red
red
maroon
Table 3: LOCAL brand lipstick examined.
Sample No.
Solvent System
of red dye concentrations in these different (well known) and (local)
brands of lipsticks, there is no characteristic data to distinguish among
the lipstick sources, and therefore the concentrations of the red dye is
not be a unique identifier for the lipstick sources. The concentration
of red pigment differs among these “well known” and “local” brands
within a small constant sample of red lipsticks.
The chromatogram of local lipsticks (Table 3) shows that majority
of the local brands contains more than 2 dyes pigments given in table
4. While branded lipsticks contains only 1 or 2 dyes pigments given in
table 5. There is no significant difference in the hRf values among the
local and well known brands of lipsticks which can be used as unique
feature.
References
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Toluene/Benzene (4:12)
2
Toluene/Benzene/Cyclohexane (4:12:4)
3
Toluene/Benzene/Diethyl ether (4:12:2)
Table 4: Mobile or Solvent phase.
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Brand
Color
1B
bridal dream 104
revlon
spunky red
2B
cherry sparkles 57
revlon
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red flame 13
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4B
red spice
street wear
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5B
crystal shine
street wear
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6B
color burst lipstick plum
revlon
maroon
Table 5: REVLON brand lipstick examined.
hRf values (7,12,27,37,47,76), sample 2L shows four pigments having
hRf values (13,27,37,76), sample 3L shows five pigments having hRf
values (3,13,25,42,79), sample 4L shows four pigments having hRf
values (3,13,37,79), sample 5L shows three pigments having hRf values
(13,36,78) and sample 6L shows five pigments having hRf values
(3,13,29,36,80). This shows that there is no significant difference in the
hRf values among the different local brands of lipsticks under study.
For well known brands
Rf =
Distance traveled by one lipstickcomponent from the origin ( column 3)
Distance the solvent moved fromthe origin ( column 4 )
Table 2 shows the result obtained from the local brands of lipsticks.
In which sample marked as 1B shows one pigment having hRf value
(4), sample 2B shows Two pigments having hRf values (3.5,13), sample
3B shows two pigments having hRf values (25,15.5), sample 4B shows
two pigments having hRf values (2,13), sample 5B shows two pigments
having hRf values (2.5,15) and sample 6B shows two pigments having
hRf values (2.5,15). This shows that there is no significant difference in
the hRf values among the different well known brands of lipsticks under
study.
Discussion and Conclusion
The conclusion is that through thin layer chromatography analysis
J Anal Bioanal Techniques
ISSN:2155-9872 JABT, an open access journal
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