This is a description of how to use paper chromatography of amino acids (to identify amino acids).

Aims

• To demonstrate separation of amino acids by paper chromatography
• To use chromatography to identify amino acids

Jump to: navigation, search In chemistry, an amino acid is any molecule that contains both amino and carboxylic acid functional groups. ... Chromatography is a family of analytical chemistry techniques for the separation of mixtures. ...

Materials

• Chromatography tank and lid
• Chromatography paper (or filter paper), to fit tank
• Capillary tubes or pasteur pipettes
• Amino acid samples in 1molar hydrochloric acid (IRRITANT)
• Unknown samples (e.g. 'unknown' mix or filtered fruit juice)
• Solvent (BAW): Butan-1-ol : Acetic acid : Water 60 : 15 : 25 (FLAMMABLE, TOXIC)
• Hairdryer or heating tray
• Spray can of Indanetrione (Ninhydrin) 1% in butan-1-ol (FLAMMABLE, TOXIC)
• Latex gloves
• Access to fume cupboard
• Drying oven at 100°C

Filter paper is any of various grades of coarse unfinished paper which can be used to separate solids from a liquid in which they are suspended. ... Capillary action or capillarity is the ability of a narrow tube to draw a liquid upwards against the force of gravity. ... Jump to: navigation, search The chemical substance hydrochloric acid is the aqueous (water-based) solution of hydrogen chloride (HCl) gas. ... Butanol is a higher alcohol with a 4 carbon atoms and a general formula of C4H10O. There are 4 different isomeric structures for butanol: butan-1-ol CH3-CH2-CH2-CH2-OH butan-2-ol CH3-CH2-CH(OH)-CH3 sec-butanol CH3-CH-CH3 | CH2OH OH | tert-butanol CH3... The chemical compound acetic acid (from the Latin word acetum, meaning vinegar), systematically called ethanoic acid, is the acid that gives vinegar its sour taste. ... The LaTeX logo, typeset with LaTeX LATEX is a document preparation system for the TeX typesetting program. ...

Methods

BAW solvent should be made up fresh on the day.
Using a pencil, lightly draw a line 1.5–2 cm above the bottom of the chromatography paper. Make small marks at 1.5 cm intervals along the line. Fill a capillary tube or pipette by capillary action with your first sample. Touch the tip of the capillary to the first mark and pull it away. You should leave a small (>5 mm) wet spot. Let this dry and apply the sample again. Make a note of which sample is placed on the spot (you can lightly write on the paper beneath the mark, using a pencil). Repeat for each sample or mixture, on a different mark and using a fresh tube or pipette each time. Let the paper dry. Capillary action or capillarity is the ability of a narrow tube to draw a liquid upwards against the force of gravity. ...

Put 1 cm or less of BAW into the chromatography tank. The sample spots should not dip into the solvent. Place the tank in a cool place out of direct sunlight. Stand the chromatography paper in the tank so that the bottom edge is in the solvent but the remaining paper does not touch the tank. Place the lid on the tank and leave for about 1 hour.

Wearing gloves, remove the damp paper and mark where the solvent has reached. Dry the paper in the fume cupboard. Still in the fume cupboard, spray the whole of the paper with ninhydrin in butanol. Dry the paper and transfer it to the drying oven for 1–2 min. A fume hood, also called a fume cupboard, or colloquially just a hood, is a critical piece of safety equipment in any chemistry laboratory, designed to ensure proper ventilation of hazardous fumes. ... Ninhydrin is a chemical used to detect ammonia or primary amines. ...

Mark the position of each spot that develops. Note the colour and Rf (or travel distance) of each standard and hence find the amino acid composition of the unknown samples.

Hazards

• Acetic acid (Ethanoic acid)
  Irritant vapour. On skin; flood with water. Ventilate area.
• Butan-1-ol
  Highly flammable. Remove ignition sources
  Harmful by inhalation. Irritant vapour. Use fume cupboard.
• Hydrochloric acid 1 molar
  Irritant, may cause burns. On skin; flood with water. Spills, neutralise with sodium bicarbonate.
• Indanetrione
  Irritant. Wear gloves. Indanetrione, spilt on the skin, is not immediately noticeable. It stains skin purple, slowly, and even seems to work through gloves.
  Wash frequently to remove it.
• Hot Oven; also produces Butan-1-ol fumes.
  Remove papers wearing gloves, or use forceps. Ventilate area

See also

• School Biology experiments and demonstrations
• How-to's