Thin Layer Chromatography at Higher and Advanced Higher Biology Summer School 2013 Thin Layer Chromatography at Higher and Advanced Higher Kath Crawford, Paul Beaumont, Lorraine Bruce

Unit: Sustainability and Interdepedence 1 The science of food production (b) Plant growth and productivity Photosynthesis Energy capture by photosynthetic pigments....

Unit: Cells and Proteins Laboratory techniques for biologists (c) Separation techniques Unit: Investigative Biology 2. Experimentation

Thin Layer Chromatography Separation of complex mixtures between two phases: a stationary phase a mobile phase

Thin Layer Chromatography In TLC: the stationary phase is a thin layer of absorbent particles, in this case silica gel the mobile phase is the solvent

Fisher Scientific UK. Tel: 01509 231166 Cat No: TLC-410-610X Price £35 for pack of 50 (5 x 20 cm)

Thin Layer Chromatography Separation of: photosynthetic pigments in leaves anthocyanins in leaves, flowers, fruit and vegetables

Method Place ca. 1 cm2 leaf in mortar [If tissue is coarse, silver sand may be added] Grind until all tissue broken up Add ca. 750 ml propanone Pour off supernatant into a container and seal

Method Mark (with a pencil) a plate 1.5 cm up from the base on the edge of both sides Use a ruler across these marks to mark two spots 2 cm apart in the middle of the plate

Method Use Pasteur pipette to place small drops of supernatant Dry off before adding another lot of drops Continue until there is a concentrated line of colour

Method Repeat process until a second line is produced

Method Plate into a 600 cm3 beaker, replace foil Remove plate when solvent nearly at top Immediately mark solvent front

Student results

Results Pigment Colour RF value carotene yellow-orange 0.91 pheophytin a grey 0.75 pheophytin b light grey 0.63-0.75 chlorophyll a blue green 0.63 chlorophyll b green 0.58 xanthophylls yellow 0.53 0.47 0.32

Coleus