Joshi et al.

, J Anal Bioanal Techniques 2013, 4:1

Analytical & Bioanalytical 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
Forensic Science, Amity Institute of Forensic Sciences (AIFS), Amity University, India
2
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 signiicant 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; • Exposing to luorescent dyes

Lipstick samples; TLC; Solvent system; hRf value
• Exposing to iodine
Introduction • Spraying with a reagent
In forensic science, comparative examinations are usually based hese procedures may be used alone or in conjunction to make the
on the physical or chemical nature of a substance, or both. Lipsticks
components of a sample visible. he distance a component has traveled
are composed of waxes, oils, organic dyes [1], and inorganic pigments
up a plate can be assigned a numerical value known as the Rf value.
[2,3]. Color matching can do identifying the lipstick responsible for
Rf is deined as the distance traveled by the component divided by the
leaving a smear. his color analysis may be used to identify the lipstick
found at the crime scene [4]. 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
he colors of lipstick are oten due to a mixture of several pigment from the chamber and dried.
compounds. hese pigments can be separated using thin layer
chromatography. Depending on the type of pigment, the mobile phase Materials and Methods
will vary. Lipsticks are soluble in toluene, so toluene serves as the mobile
Jar with lid, lipstick samples (Branded and Local), TLC plates,
phase. Ater separation, the chromatogram is complete and illustrates
mobile phase/solvent phase, clean cotton piece, sterilized scissor, ine
the diferent pigments that make up a particular color of lipstick [2,3,5-
capillaries, ruler, pencil, todine pellets, titer plates, graduated cylinder,
10].
gloves, protective mask and thirty samples of lip impressions bearing
Traces of Lipsticks, cosmetics, nail polish, or other smears could be lipstick were obtained on cloth using various brands and types of
found let on drinking cups, glasses, cigarette butts, and tissue papers lipsticks, which were available from volunteers [5,13,18-20].
and may all be signiicant forensic evidence [11] in the investigation of
a crime, especially in cases such as a sexual assault or a homicide [12].
hin-Layer Chromatography (TLC)
his physical evidence may be found on clothing, parts of the body, a A thin-layer plate is prepared by coating a glass plate with a thin
tissue, or cigarette [5,13]. By comparing the composition of a lipstick ilm of a granular material. Commonly, silica gel or aluminum oxide or
smear with that of a victim, forensic scientists can demonstrate indirect cellulose immobilized onto a lat, inert carrier sheet is used, but paper
proof of contact or a relationship between victim and suspect. Also, it is may suice in simpler experiments. hese serve as the solid stationary
sometimes possible to extract saliva DNA from the print and may link phase. If the sample to be analyzed is a solid, it must irst be dissolved
a suspect to a crime scene [2]. in a suitable solvent. A few micro liters of the solution are then applied

Various methods of forensic lipstick analysis were reported [6,8,14-

17]. Small amount of lipstick (approximately 10 µg) could lead to good *Corresponding author: Kapil Verma, M.Sc Forensic Science, Amity Institute
comparisons in TLC [3]. hin Layer Chromatography (TLC) is a widely- of Forensic Sciences (AIFS), B-Block, Lower Ground Floor, Amity University,
used chromatography technique used to separate chemical compounds. Uttar Pradesh, Sec-125, Noida-201303, Uttar Pradesh, India, E-mail:
forensic.kapilalert@gmail.com
hin-layer chromatography incorporates a solid stationary phase and
Received November 22, 2012; Accepted December 17, 2012; Published
a moving liquid phase to cause a separation of the constituents of a
December 24, 2012
mixture. Although simple test may be run by simply allowing a solvent
Citation: Joshi B, Verma K, Singh J (2013) A Comparison of Red Pigments in
to pick up a piece of porous paper, a more revealing test requires the Different Lipsticks Using Thin Layer Chromatography (TLC). J Anal Bioanal
preparation of a plate. Because most compounds are colorless, no Techniques 4: 157. doi:10.4172/2155-9872.1000157
separation will be noticed ater development unless the materials are Copyright: © 2013 Joshi B, et al. This is an open-access article distributed under
visualized. his is may be done by: the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and
• Exposing to UV light source are credited.

J Anal Bioanal Techniques

ISSN:2155-9872 JABT, an open access journal 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 from the spotted pencil line. Some lipsticks have only two or three
to the solid state in the same manner. he plate or paper is then placed components, and some have more. Enter these measurements on the
upright into a closed chamber that contains a selected solvent. Data Table.
he solvent will slowly begin to pick up the plate by capillary action. 13. Determine the Rf for each lipstick component of all lipstick
It is the rising solvent that serves as the moving phase in thin-layer samples. Enter those values on the Data Table. To calculate the Rf value,
chromatography. As it moves past the sample spot, the components of divide the distance traveled by each lipstick component by the distance
the sample will become distributed between the stationary solid phase traveled by the solvent.
and the moving liquid phase. hose components with the greatest
Distance traveled by one lipstickcomponent from the spotted pencil line
ainity for the moving phase will travel up the plate at a faster speed Rf =
Distance the solvent moved from thespotted pencil
as compared to those that have greater ainity for the stationary phase.
hin-Layer Chromatography (TLC) is a method use to separate Samples are run for both the samples of branded and local lipstick.
components from each other in a mixture. It is normally experimented Use hood if possible. Self life is about one month.
on glass, aluminum foil or plastic which is coated by some kind of Steps followed are shown in igures 1-11.
absorbent material (ex: silica gel, aluminum oxide). It takes the Rf
values of each color component and compares them; Rf values are Results
experiments that depend on the polarity of the substance on the paper
he data collected from the chromatograms were recorded, and
chromatography [21];
1. A piece of cotton cloth was washed in a detergent solution,
immersed in a hot water, and dried.
2. his cloth was cut into small pieces (2 cm×2 cm) and lipstick
smears were rubbed onto these pieces.
3. he stained areas from the cloth pieces were cut and transferred
into serially marked small bowls.
4. hese samples were mixed with an extracting solution and each
bowl was then shaken for about 5-10 minutes to remove the stain from Figure 1: Sample of different lipstick for study.
the cloth piece.
5. Cloth piece is taken out, and the extract was used for further
examination with TLC.
6. Obtain jar with lid, piece of ilter 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,
Figure 2: Samples rubbed on a clean cotton piece Bowls
6B for six diferent well known brands of red lipsticks on a single TLC
contains the extracted sample.
plate. Similarly 1L, 2L, 3L, 4L, 5L, 6L for six diferent local brands of
red lipsticks.
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. he 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. he mobile phase/solvent should be about 0.5 cm
deep.
Figure 3: Picture showing TLC plate.
10. Carefully insert the TLC plate into the jar, sample end down.
he lipstick dots must be above the mobile phase/solvent. Secure the
lid.
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 Figure 4: Extract preparation in Toluene (Bowls contains the
measure the distance in cm each component of the lipsticks moved extracted sample).

J Anal Bioanal Techniques

ISSN:2155-9872 JABT, an open access journal 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 10: Chromatogram of local lipsticks (Spots seen for local brand of
Figure 5: Extract preparation in Toluene (Bowls contains the extracted lipstick when viewed under Iodine fumes).
sample).
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.

Column 3 Column 4
Lipstick No. of Distance Lipstick Distance hRf Values for
Samples Spots Components Mobile Phase Each Colored
Solvent System:- Moved (cm) Moved (cm) Component
Toluene /Acetone 1L 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
2L 1 1.3 10 13
2 2.7 10 27
3 3.7 10 37
4 7.6 10 76
3L 1 0.3 10 3
Figure 7: Jar containing iodine pellets. 2 1.3 10 13
3 2.5 10 25
4 4.2 10 42
Solvent System:- 5 7.9 10 79
Toluene/Benzene/Diethyl ether 4L 1 0.3 10 3
2 1.3 10 13
3 3.7 10 37
4 7.9 10 79
5L 1 1.3 10 13
2 3.6 10 36
3 7.8 10 78
6L 1 0.3 10 3
2 1.3 10 13
3 2.9 10 29
4 3.6 10 36
Figure 8: Jar containing iodine pellets.
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.
he 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 Table 1 shows the result obtained from the local brands of
(Spots seen for local brand of lipstick when viewed under Iodine fumes). lipsticks. In which sample marked as 1L shows six pigments having

J Anal Bioanal Techniques

ISSN:2155-9872 JABT, an open access journal 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 Column 4 of red dye concentrations in these diferent (well known) and (local)
Lipstick No. of Distance Lipstick Distance Rf Values for Each brands of lipsticks, there is no characteristic data to distinguish among
Samples Spots Components Mobile Phase Colored Component the lipstick sources, and therefore the concentrations of the red dye is
Moved (cm) Moved (cm) not be a unique identiier for the lipstick sources. he concentration
1B 1 0.4 10 4 of red pigment difers among these “well known” and “local” brands
2B 1 0.35 10 3.5 within a small constant sample of red lipsticks.
2 1.3 10 13
3B 1 0.25 10 25 he chromatogram of local lipsticks (Table 3) shows that majority
2 1.55 10 15.5 of the local brands contains more than 2 dyes pigments given in table
4B 1 0.2 10 2 4. While branded lipsticks contains only 1 or 2 dyes pigments given in
2 1.3 10 13 table 5. here is no signiicant diference in the hRf values among the
5B 1 0.25 10 2.5 local and well known brands of lipsticks which can be used as unique
2 1.5 10 15 feature.
6B 1 0.25 10 2.5
2 1.5 10 15 References
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