1. Peptides Martina Zlámalová

3. Peptide Synthesis Methodology is carried out by FMOC chemistry, using PEG-Polystyrene resins peptides are cleaved from the resin and de-protected using cleavage cocktail peptides are precipitated from the cocktail using cold diethyl ether washed three times with the cold ether and then dissolved in a crude buffer

4. Reversed phase chromatography Peptides QC is based on reversed phase chromatography on a Supelco Bio Wide Pore column and by MALDI-TOF mass spectrometry. Peptides not meeting the purity criteria are purified by reversed phase chromatography.

5. Peptide Synthesis Monitoring reagents Colour test of amino acids in SPPS and SPOS Reaction of ninhydrin with free primary amines (intensive blue) Test is applied quantitatively and qualitatively Important to state – NOT yield the typicaly dark blue with serine, asparagine, aspartic acid and proline

6. Structure The test kit contains 50 ml each of the following solutions: Phenol, ~ 80% in ethanol, KCN in H 2 O/pyridin, Ninhydrin 6% in ethanol

7. Procedure 1. Remove a few resin beads from the reaction vessel and wash 3 times with ethanol 2. Transfer beads into a small glass test tube and add 3 drops of each solution 3. Mix well and heat the test- tube at 120° for 5 minutes

8. The resin beads and the solution turn dark blue when a free primary amine is present. The resin beads remain their colour and the solution stays yellow when no free primary amine is present (expected result after successful coupling). A recoupling step is necessary when a slight blue colour is detected in the solution and/or on beads

9. Thank You for Your attention!