Unit 2- Practical Scientific Procedures and Techniques
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Distinction grade assignment 2C
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Unit 2- Practical Scientific Procedures and Techniques
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PEARSON (PEARSON)
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Unit 2- Practical Scientific Procedures and Techniques
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(1) Class notes
(2) Separation by Chromatography - GeeksforGeeks
(3) https://chrominfo.blogspot.com/2020/05/factors-affecting-separation-in-
paper.html
(4) What is Chromatography? Its Definition, Principle & Uses (studyread.com)
(5) https://www.news-medical.net/life-sciences/Thin-Layer-Chromatography.aspx
(6) https://www.chemguide.co.uk/analysis/chromatography/thinlayer.html#top
(7) https://whatishplc.com/faq-hplc/how-does-temperature-affect-paper-
chromatography/
(8) https://prezi.com/apnhflnwy_ea/paper-chromatography-lab/
(9) https://www.studocu.com/en-gb/document/university-of-derby/foundation-
biology/separation-of-amino-acids-using-paper-chromatography/20273995
(10) http://faculty.buffalostate.edu/wadswogj/courses/BIO211%20Page/
lectures/lab%20pdf%27s/Amino%20Acid%20lab.pdf
Chromatography is a practical technique which can be used to find and separate
the different components in a mixture, chromatography involves a mixture which
is dissolved in a mobile phase which could be either a liquid or a gas and then it
would be passed through an immobile stationary phase which is normally a solid.
Thin layer chromatography/TLC
Grey/light green-carotin
Dark green-Phaeophytin
Yellow-Chlorophyll A
Thin layer chromatography is used to separate the compounds in non-volatile
mixtures, it is used for both quantitative and qualitative sample analysis. TLC
can be used in forensic studies and analysis where bodily fluids such as blood or
urine can be tested for drug presence; acidic and neutral drugs such as
ibuprofen and phenytoin can be identified by using octadecyl silica while in the
stationary phase, and basic drugs such as ketamine and methadone can be
identified by using plain silica and octadecyl.
, To perform thin layer chromatography, you would need to Prepare a silica gel
chromatography plate by drawing a pencil line carefully about 1.5cm from the
bottom of the slide. Then you would mark a small pencil cross at the centre of
the line. Avoid touching the surface of the plate with your fingers because this
can cause fingerprints on the sample and reduce the accuracy. Then Use a fine
glass capillary tube to transfer the pigment to the cross. Briefly touch the
surface of the plate with the tube and allow to dry. Repeat this process several
times to produce a small, concentrated spot of pigment. Now, Stand the plate in
a Coplin jar holding the solvent and replace the lid. The level of the solvent must
be below the level of the pigment spot. Allow the solvent to rise until the solvent
front reaches about 1cm from the top of the plate. Remove your completed
chromatogram, mark the solvent front and allow to dry. Then, using a pencil,
circle the spot then finally Calculate the Rf values for each spot. (1)
Paper chromatography with plant pigments.
Rf value = 0.883
5.3/6=0.883
The mobile phase in this type of chromatography is the plant pigments and the
toluene, and the stationary phase is the plant pigments. To do paper
chromatography you would have to put a drop of a solution of a mixture of dyes
or inks onto a piece of chromatography paper and allowed to dry. The mixture
will begin to separate as the solvent front advances past the mixture. Simple
materials such as ink front fountain and fibre tip pens, food colouring and dyes
can be separated using this type of chromatography. First you need to ensure
you’re wearing the correct PPE such as goggles, gloves and a lab coat. To
prepare the sample you need to take a strip of chromatography paper which is
around 19cm long and use a sharp pencil to draw a line around 2cm away from
the end of the paper, dab your plant pigment along the line ensuring that you
fully cover the line and then leave to dry for a few minutes. Then place the
sample into the Coplin jar containing toluene and allow them to separate, once
the solvent front is about 2/3 way up the paper, remove it and mark the solvent
front line with a pencil and allow the completely dry. Once dry you can calculate
the rf value by doing the distance pigment travels / distance solvent front
travels. (1)
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