KROMATOGRAFI LAPIS TIPIS

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Kromatografi Lapis Tipis (KLT)

Kromatografi lapis tipis (KLT) adalah teknik

kromatografi yang digunakan untuk
pemisahan dengan fasa diam padat dan
fasa gerak cair.
Fasa diam berupa zat padat (adsorben)
yang dilapiskan pada lempeng kaca,
plastik atau aluminium setebal 0,1-0,3 mm.
Fasa diam berupa silika , alumina atau
selulosa
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Lempeng yang lazim djual berukuran

20x20 cm.
Karena kesederhanaan & kecepatan
KLT, seringkali digunakan untuk
memonitor perkembangan reaksi
organik dan mengecek kemurnian
produk

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Stationary Phase: Silica (SiO2)

OH
Si
O

Si
O
O
Si

OH

OH

Si
O
O
Si

O
O
O

Si
O
O
Si

O
O
Si

OH

OH
O
O
Si
O
O
Si

O
O

Si

O
O

O
O

O
O
Si

O
O

THIN LAYER CHROMATOGRAPHY

Individual components move up at different rates,
depending on intermolecular forces between the
component and the silica gel stationary phase and
the component and the mobile phase.

The stationary phase is SiO2 and is very polar.

http://www.instructables.com/id/EW1YDCYF4REC0IU/

It is capable of strong dipole-dipole and H-bond donating and accepting

interactions with the analytes (the components being analyzed).
More polar analytes interact more strongly with the stationary phase in move
very slowly up the TLC plate.
By comparison, the mobile phase is relatively nonpolar and is capable
of interacting with analytes by stronger London forces, as well as by dipoledipole and H-bonding.
More nonpolar analytes interact less strongly with the polar silica gel and more
strongly with the less polar mobile phase and move higher up the TLC plate.

Stationary Phase: Alumina

OH

OH

OH

OH

Al

Al

Al

Al

Al

Acidic: -Al-OH
Neutral: -Al-OH + -Al-OBasic: -Al-O-

TLC Stationary Phases

www.vwr.com

www.vwr.com

Adsorption of Solutes
The adsorption strength of compounds increases with increasing polarity of
functional groups, as shown below:
-CH=CH2, -X, -OR, -CHO, -CO2R, -NR2, -NH2, -OH, -CONR2, -CO2H.
(weakly adsorbed)
(strongly adsorbed)
(nonpolar)
(more polar)

Elution Strength of Mobile Phase (

Elution strength is generally considered to be equivalent to polarity. A solvents

elution strength depends on Intermolecular Forces between the solvent and the
analytes and between the solvent and the stationary phase.
A more polar (or more strongly eluting solvent) will move all of the analytes to a
greater extent, than a less polar, weakly elution solvent.
For example, the elution strength of hexane is very low;
the elution strength of ethyl acetate is higher;
the elution strength of ethanol is even higher;

= 0.01.
= 0.45
= 0.68

Solvent Properties and Elution Strengths

IDENTIFIKASI ZAT
LAZIMNYA DIGUNAKAN HARGA Rf
MESKIPUN KURANG TEPAT ( bila
dibandingkan pada
kromatografikertas)
Lebih baik menggunakan penampak
noda
Harga Rf senyawa senyawa murni
dapat dibandingkan dengan senyawa
standar
Rf = jarak yang digerakkan senyawa
dari ttk
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awal

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Harga Rf hanya berlaku utk campuran

tertentu dari pelarut & penyerap yang
digunakan
Identifikasi lainnya dengan Rx atau
Rstd
Rx = Jarak yang digerakkan senyawa yang tidak diketahui
Jarak yang digerakkan senyawa standar yang
diketahui

Senyawa standar biasanya memiliki

sifat kimia mirip dengan senyawa
yang dipisahkan pada kromatogram
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FAKTOR YANG MEMPENGARUHI

GERAKAN NODA / Rf
Struktur kimia senyawa yang
dipisahkan
Sifat penyerap & derajat aktifitasnya
( pemanasan dalam oven)
Tebal & kerataan lapisan penyerap
(fasa diam)
Pelarut & derajat kemurnian fasa
gerak
derajat kejenuhan uap dalam bejana
pengembang
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Teknik percobaan (penurunan ,

penaikan & mendatar) meskipun
penurunan merupakan teknik yang
paling umum digunakan
Jumlah sampel yang digunakan
(penotolan cuplikan berlebihan dapat
menimbulkan tailing & Rf sulit
ditentukan)
Suhu ( sebaiknya pada suhu tetap)
Kesetimbangan ( dalam bejana
pengembang pelu jenuh dengan uap
pelarut)
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THIN LAYER CHROMATOGRAPHY

Calculation of Rfs
2.0 cm
5.0 cm

= 0.40

Rf (B) = 3.0 cm

= 0.60

Rf (A) =
Solvent Front

5.0 cm

Distance solvent
migrated = 5.0 cm
4.0 cm
Distance A
migrated = 3.0 cm

5.0 cm

Distance B
migrated = 2.0 cm

Distance C
migrated = 0.8 cm
Origen

Rf (C) = 0.8 cm = 0.16

x
A

x x x x
B U C D

3.0 cm

Rf (D) = 4.0 cm = 0.80

0.8 cm

Rf (U1) = 3.0 cm = 0.60

5.0 cm

5.0 cm

Rf (U2) =

0.8 cm
5.0 cm

= 0.16

The Rf is defined as the distance the center of the spot moved divided
by the distance the solvent front moved (both measured from the origin)

Resolution

The separation between two analytes on a

chromatogram can be expressed as the
resolution, Rs and can be determined using
the following equation:
Rs = (distance between center of spots)
(average diameter of spots)
In TLC, if the Rs value is greater than 1.0, the
analytes are considered to be resolved.
x

Improving Resolution:

For two closely migrating components, optimum resolutions

are usually obtained when the Rfs of both compounds
are between 0.2 and 0.5
* To Improve Rs, change the elution strength of the solvent
to optimize Rfs

change onet (= in capacity factor), all compounds will

be effected similarly.

Alter the composition of the solvent system so that the

components affinity for the mobile phase vs. the solid
phase are differentially changed (= change in
selectivity).

Changing the chemical nature of the solvent system,

such as changing a hydrogen bonding solvent to a
solvent which cannot hydrogen bond to the analyte, is
often the most effective.

** Improve Rs by decreasing the diameter of the

analyte spots. This can be achieved by applying
smaller and less concentrated spots.

http://orgchem.colorado.edu/hndbksupport/
TLC/TLCprocedure.html

Metoda

Sampel, baik sebagai larutan atau

dilarutkan dalam pelarut yang volatile ,
didepositkan sebagai suatu noda pada
permukaan fasa diam.
Konstituen sampel yang belum diketahui
dapat diidentifikasi dengan mengelusinya
secara simultan bersama-sama dengan
standar yang sudah diketahui

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Ujung bawah plat diletakkan dalam pelarut,

kemudian pelarut dijalankan di atas permukaan
plat dengan gaya kapiler.
Pada saat bagian depan pelarut mencapai ujung
lain fasa diam, plat dikeluarkan dari fasa pelarut.
Noda-noda
yang
terpisah
dimunculkan
menggunakan sinar
ultraviolet atau menempatkan plat dalam uap iodium.
Komponen yang berbeda-beda dalan capuran
bergerak di permukaan plat dengan kecepatan
berbeda-beda karena perbedaan sifat partisinya
diantara fasa cair yang mobil dan fasa diamnya.
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Spotting a TLC plate with

sample:

Running the TLC plate in

solvent:

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KULIAH KA II-A

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KLT 2 DIMENSI
Two-dimensional TLC uses the TLC
method twice to separate spots that are
unresolved by only one solvent
After running a sample in one solvent, the
TLC plate is removed, dried, rotated 90 o,
and run in another solvent.
Any of the spots from the first run that
contain mixtures can now be separated.
The finished chromatogram is a twodimensional array of spots.
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KLT Dua Dimensi

KULIAH KA II-A

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KLT PREPARATIF

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