Identification of Rhodamine B in

Lipstick by TLC Method and UV-VIS

Spectrophotometry
GROUP 2:

1. AMBARWATI
2. LUSI FEBRIANITA KOBANDAHA
3. NENI PURNAMA DJAPANO
4. SITI ROMLA MASPEKE
 THE SUBJECT

I INTRODUCTION R RESEARCH METHODS

R RESULTS & DISCUSSION C CONCLUSION

 #01
INTRODUCTION
Rhodamine B is a synthetic dye that is
generally used as a dye for paper and
textiles. The use of Rhodamine B as a
cosmetic dye can cause skin irritation,
eye irritation and is carcinogenic.
Given these dangers, the use of
Rhodamine B dye can be detrimental
and endanger public health
Identification of Rhodamine B in
Lipstick by TLC Method and UV-VIS
Spectrophotometry
Lipstick is a cosmetic
preparation made of wax,
pigment or dye and oil. Dyes
have a role in lipstick colouring
so that the resulting lipstick
gives a very attractive colour
and appearance so that
consumers are interested in
using it.
 #02
RESEARCH METHODS
 TOOL AND MATERIALS

Tools Materials

- drop pipette - Analytical balance • Lipstick

- separating funnel - Measuring cup • hydrochloric acid

- filter paper - Beaker • Methanol

- thin layer chromatography - water bath • Ether

- UV-Vis spectrophotometer - spatula • comparison standard (BP)

for Rhodamine B dye.
 RESEARCH PROCEDURE
11 Lipstick samples were taken from large markets in East Jakarta, namely the
Karamat Teak Market, Jatinegara Market, Cakung Market and Old Klender Market.
1. Preparation
 Test solution
An amount of approximately 500 mg of the sample was added with 4 drops of
4 M hydrochloric acid, added 5 ml of methanol, then melted in a water bath,
then added methanol to 10 ml and filtered through filter paper (A). 
 Comparative standard solution
50 mg of Rhodamine B dye as reference standard was dissolved in 100 ml of
methanol (B).
 A mixture of the test solution and reference standard
A mixture of solutions (A) and (B) with the same volume is made to produce a
solution (C).
2. Implementation and Observation
a). How to identify with thin layer chromatography
Solutions A, B and C were each spotted separately and thin-layer chromatography was
performed as follows:
- Stationary phase: silica gel G
- mobile phase:
i. Ethyl acetate – n – butanol – ammonia (20 : 55 : 25)
ii. Ethyl acetate – methanol – ammonia (15 : 6 : 3)
iii. n - propanol - ammonia (90 : 10)
- Saturation: with filter paper
- Volume of spotting : Solutions A, B and C each 10 L Propagation distance 12 cm
- Spot viewer:
i. No visible spots, red spots
ii. with UV 254, the spots fluoresce yellow.
b). How to identify with UV-Vis Spectrophotometry
 Test solution
Weighing approximately 5 g of the sample was put into a beaker glass, then added 30 ml of 2% sodium
hydroxide solution, stirred and heated over a water bath until it melted. The liquid was put into a 100 ml
separatory funnel, then added 30 ml of ether, shaken for 3 minutes and allowed to stand to separate. Then
the aqueous phase was removed, the ether phase was washed twice with 20 ml of 0.5% sodium hydroxide
solution. Then the washing was removed, the ether phase was added to 10 ml of 0.1 N hydrochloric acid
and shaken, the acid phase was accommodated (A).
 Comparative standard solution
Weighing approximately 50 mg of Rhodamine B as a reference standard dissolved in 250 ml of methanol.
Then an amount of about 1 ml of this solution is added with 25 ml of 0.1 N hydrochloric acid (B).
 A mixture of the test solution and reference standard
A mixture of solutions (A) and (B) with the same volume is made to produce a solution (C).
 Identification
The maximum absorption of solutions A, B and C were each measured at a wavelength of approximately
558 nm using 0.1 N hydrochloric acids as a blank.
 #03
RESULTS & DISCUSSION
 The data obtained from testing using thin layer chromatography are as follows:

Table 1. Rf value data for samples in 3 kinds of eluents.

No Rf Value
Table 2. Wavelength data (λ ) maximum
Eluen A B C
No Description λ Maximum Absorbance
BP. Rhodamin B 0,9 0,91 0,9
1. Sampel 1 0 0 0
1. BP Rhodamin B 558 0,7579
2. Sampel 2 0 0 0
3. Sampel 3 0 0 0
2. Sampel 4 557 0,3120
4. Sampel 4 0,85 0,95 0,8
5. Sampel 5 0 0 0
6. Sampel 6 0 0 0
7. Sampel 7 0 0 0
The results of the above thin layer chromatography
8. Sampel 8 0 0 0
obtained 1 positive sample containing Rhodamine B then
9. Sampel 9 0 0 0
tested using UV-Vis Spectrophotometry.
10. Sampel 10 0 0 0
11. Sampel 11 0 0 0

The process for separating the
dye contained in the sample is by
dissolving the sample using methanol.
The dye dissolved in methanol was
then identified using thin layer
chromatography and further
identification using a UV-Vis
spectrophotometer for a confirmation
test. In this study, thin layer
chromatography was used for the
preliminary test, then confirmed by
using the spectrophotometric method
to ensure a true positive result for
rhodamine B.
Discussion
The sample was prepared
before hand to remove the
nterfering substances contained in it
to facilitate the dye extraction
process. The next step is to dissolve
the sample with methanol. After
identification using thin-layer
chromatography of 11 samples, 1
sample containing Rhodamine B
dye was obtained.
Some samples were positive for
rhodamine B using either the TLC
or spectrophotometric methods,
meaning that there were still people
who abused the use of rhodamine B
as an electrical dye. This will
endanger the user because
continuous use can cause skin
irritation, even cancer and tumours.
used UV-Vis spectrophotometry to
prove that the sample was positive
for Rhodamine B.
 #04
CONCLUSION
 Of the 11 lipstick samples circulating in four
markets in East Jakarta, it was identified that 1
sample (9.090% of 11 samples) contained
Rhodamine B dye.
THANKS