FRS 581: FORENSIC CHEMISTRY

EXPERIMENT 4:

TLC OF INKS

NAME: NURUL HIDAYAH BT HASNI @ MOHD ZIKRI

MATRIX NO: 2019726077

GROUP: AS2535B

LECTURER’S NAME: MOHAMED IZZHARIF ABDUL HALIM

DATE OF SUBMISSION: 26 NOVEMBER 2020

EXPERIMENT 4: TLC OF INKS

OBJECTIVE

i) Understanding and experiencing ink analysis using Thin-layer chromatography

technique.
ii) Determining the retention factor of ink from ball point pen, gel pen, and roller ball pen.
iii) Determining the difference and similarities of characteristic of all those ink samples.

PROCEDURE

1) Mobile phase was prepared with 7.5 mL ethyl acetate, 3.5 mL 95% ethanol and 3.0 mL
distilled water was added into a 100 mL beaker and was sealed tightly with parafilm.
2) Filter paper was placed into the beaker to saturate the mobile phase vapor in the beaker.
3) Then, forged documents of handwriting were made using all the ink samples (ball point
pen, gel pen, and roller ball pen).
4) The forged documents then were crumpled and placed in a test tube with two drops of
95% ethanol. The test tube was swirled gently.
5) The solubility of the ink samples was observed.
6) A line was marked at the bottom of a short edge of the TLC plate by using a pencil.
7) The TLC plate was placed in the mobile phase beaker immediately and sealed off tightly.
8) The TLC plate was removed when the solvent front reached about ¾ of the TLC plate
and allowed to dry.
9) The Video Spectral Comparator 500 was used to observe the TLC plate under normal
light, Fluorescence and Ultraviolet with different wavelength 254 nm, 312 nm, and 365
nm.
10) The Rf value of each band was calculated and compared.
RESULTS AND OBSERVATION

Ink Samples Description of Ink

a) Roller Ball Pen • Faber Castle CX Plus FC530A9C
• Black color
• Dye base ink

Figure 1: Image of Roller Ball Pen used

b) Gel Pen • Pilot
• BL WG 5 B
• Black color
Figure 2: Image of Gel Pen used
• Pigment base ink
c) Ballpoint Pen (Black) • Pentel 0.7 (F) BXS 7E
• India
• 1809 14
• Black color
• Dye base ink

Figure 3: Image of Black Ballpoint Pen used

d) Ballpoint Pen (Blue) • Blue color
• Pilot BP-1 RT M 1710
• Dye base ink

Figure 4: Image of Blue Ballpoint Pen used

Table 1: Ink samples and description of each sample.
A. Forged Document

Ink Sample Images

Roller Ball Pen

Figure 5: Handwriting using Roller Ball Pen

Gel Pen

Figure 6: Handwriting using Gel Pen

Ballpoint Pen (Black)

Figure 7: Handwriting using Black Ballpoint Pen

Ballpoint Pen (Blue)

Figure 8: Handwriting using Blue Ballpoint Pen

Table 2: Forged handwriting using each ink sample.
B. Thin-Layer Chromatography Paper After Removed from Mobile Phase Chamber

Figure 9: Spot produced by each of ink samples on TLC paper

C. Normal Light

Figure 10: Appearance of each ink sample’s spot under normal light
D. Fluorescence

Figure 11: Appearance of each ink sample’s spot under fluorescence light
E. Ultraviolet
Wavelength (nm) Result

254

Figure 12: Appearance of each ink sample’s spot under

UV-light at 254nm

312

Figure 13: Appearance of each ink sample’s spot under

UV-light at 312nm
 365

Figure 14: Appearance of each ink sample’s spot under

UV-light at 365nm
Table 3: Observation of each ink sample under UV light

F. Solubility Test
Samples Solubility Observation
Ballpoint Pen (Black) Soluble Solution turn violet
Roller Ball Pen (Black) Soluble Solution turn violet
Gel Pen (Black) Not Soluble No changes
Ballpoint Pen (Blue) Soluble Solution turn blue color
Table 4: Solubility test observation for each of ink sample.

Figure 15: Observation for each of ink sample undergoes solubility test
G. Calculation

𝐷𝑖𝑠tan𝑐𝑒 𝑡𝑟𝑎𝑣𝑒𝑙𝑙𝑒𝑑 𝑏𝑦 𝑐𝑜𝑚𝑝𝑜𝑛𝑒𝑛𝑡

Formula of Retention factor value, Rf = 𝐷𝑖𝑠tan𝑐𝑒 𝑡𝑟𝑎𝑣𝑒𝑙𝑙𝑒𝑑 𝑏𝑦 𝑠𝑜𝑙𝑣𝑒𝑛𝑡 𝑓𝑟𝑜𝑛𝑡

Roller Ball (Black) Ballpoint (Black) Ballpoint (Blue)

Spot 1 Spot 1 Spot 1
2.6 𝑐𝑚 2.8 𝑐𝑚 2.8 𝑐𝑚
Rf = 9 𝑐𝑚
Rf = 9 𝑐𝑚
Rf = 9 𝑐𝑚

= 0.290 = 0.311 = 0.311

Spot 2 Spot 2 Spot 2

4.0 𝑐𝑚 3.9 𝑐𝑚 3.5 𝑐𝑚
Rf = 9 𝑐𝑚
Rf = 9 𝑐𝑚
Rf = 9 𝑐𝑚

= 0.440 = 0.433 = 0.390

Table 5: Calculation to determine the Rf value of each ink sample.
DISCUSSION

In forensic laboratory, thin-layer chromatography (TLC) is one of the most common

chromatographic methods and one of the simplest techniques in separation chemistry. In this
method, the stationary phase is a thin layer pf gel adsorbent coated onto plastic or metal backing,
which is the TLC plate. By capillary action, the liquid mobile phase that is drawn into the gel
will carried up toward the top of plate. Along the way, the samples are separated into their
constituent chemicals. While it has many applications in many industries, in forensic labs, it
helps the scientist to determine if two pieces of text or signature were written using the same pen
or ink, which can indicate fraud or forgery.

In this experiment four types of pen were used to study analysis of ink using TLC. For the first
part, the appearance of the signature using each of the ink types were observed and the images
were captured. Through naked eye, all the signature looks the same except for the color of the
sample using blue ballpoint pen as other pen used were in black. It can also be seen that the
breadth of the signature using gel pen was a bit thick compared to other ink types. Next, the ink
samples were put in water to observe their solubility. According to Table 4, only the Gel Pen’s
ink was not soluble in water as there is no changes of color unlike the other ink samples. This is
because the ink base of gel pen is made up of pigment base while the other sample are made up
of dye base. The difference between these two bases is that dye ink uses color substances that are
dissolved in a liquid while pigment ink uses small color particles that are held in a suspension. A
pigment ink, when applied to the surface of the paper, it will form a coating over the paper and
the ink are only physically bound, not chemically. The dye ink, on the other hand, will
chemically bound to the paper. Therefore, only ink from Roller Ball Pen and Ballpoint Pen
produced spot or band on the TLC paper after it have been removed from the chamber as shown
in Figure 9.
After the TLC paper have been removed from the chamber, we can see separated spot on the
paper for sample from Roller Ball Pen and each of Ballpoint Pen. The spots were produced due
to the separated components of the ink during capillary action. The solvent assists in carrying the
separated components of the ink (solute) and because each solute have different affinities toward
the mobile phase, it will subsequently move further up the TLC paper. As the separation pattern
of the component dyes is distinctly different for each ink sample compositions, many points of
comparison can be known between each ink sample. Solutes that spend more time in a stationary
phase will be seen by the spot on the TLC paper that is closer to the starting point. Based on this
theory, it can be said that roller ball pen spends more time on the stationary phase as shown on
Figure 9. Next, the TLC paper was also observed under different light environment to further
study the difference of each ink samples.

Under normal light, the color of each spot produced are different for different type of pen. As
shown in Figure 10, spots produced by roller ball pen and black ballpoint pen give bluish-purple
and yellow spots meanwhile blue ballpoint pen produced purple and blue spots. The length of
each spots was also different for each type of sample. The length of roller ball pen ink sample
spots was more elongated compared to other two sample. This can be due to the different
components in the ink base. Furthermore, the appearance of each sample’s spot was observed
under fluorescence light. The ink of each type of sample fluoresces white under the light indicate
that some components made up the ink positively react to the fluorescence light. The length of
each spots was also different when observed under this light. Lastly, the TLC paper was also
observed under UV light. Between all the wavelength of UV light applied for the observation,
wavelength at 312nm shows the clearest result. Under this light, the distance and the number of
each spot was clearly seen, making it easier for ink distinction and distance calculation.

For the last part of experiment, the Rf value for each marked positions of noticeable components
were calculated. Based on Table 5, the Rf value are different for each of the sample.
Identification of each ink sample relies on comparing it with number of Rf literature values. Note
that even for the same type of pen (ballpoint pen), the calculated value is still different. As such,
we can conclude that, each dye components and constituents of an ink are distinctly different
from each other.
CONCLUSION

In conclusion, each type of sample shows different type of reaction when undergoes
different type of analysis. The difference and similarities of each inks were determined, and the
retention factor were also successfully calculated. The analysis of inks using TLC method was
successfully applied and done.

QUESTION

1. Apart from TLC, what other methods can be used to differentiate ink?
Other methods that can be use aside from TLC are gas chromatography (GC) and liquid
chromatography (LC). Both methods have been successfully employed in the analysis
and comparison of inks. In addition, infrared spectrophotometer also can be use for
characterizing inks by showing the absorption peaks of all the components in the ink at
one time, including the solvent, dyes, and additives.

2. How does ink analysis helps in forensic investigation?

In forensic investigation, identifying the type of ink used in a questioned document can
determine the origin of the ink. Thus, can be useful to detect alteration on a document.
Ink used on a document can look visually similar, but further investigation can show that
they are chemically different indicates that it was not included at the time of its original
writing.

3. What are the advantages and disadvantages of using TLC in ink analysis?
The advantages of TLC are that they are very simple to use and inexpensive.
Furthermore, less materials are needed for TLC which are the chamber, watch glass,
capillary, plate, solvent, pencil, and UV-light. In addition, more than 1 compound can be
separated on a TLC plate as long as the mobile phase used is preferred for each
compound. Using TLC, identification of most compound can be done by simply checking
the Rf literature values.

The disadvantages of TLC are that they do not have long stationary phase and their
detection limit is a lot higher compared to other methods. Having a short stationary phase
makes the length of separation is limited compared to other chromatographic techniques.
Lastly, TLC operate as an open system, thus factor such as humidity and temperature can
consequences to the results of chromatogram.