Final Sequence: Ac-Gly-Tyr-Arg
Peptide [M+H]+ ion
mg peptide: 6.6 mg .... Peptide: 436.21 mg
n(Resin)= 0.05 mmoles (MW = 460). You'll need this for your determination of limiting reagent, theoretical yield and %yield.
Method:
Fmoc-Arg(Pbf)-Rink Resin is supplied in a filtered syringe which is the reaction vessel for this SPPS process. Solvents and reagent solutions are added through the top of the syringe and then drained through the filter. The resin is suspended in DMF for 30 minutes to ensure complete resin swelling.Deprotection: The solvent was decanted off and replaced by a 20% piperidine in DMF solution (5mL). After sitting for 10 minutes with occasional stirring, the solvent was again decanted off and the resin washed with DMF (5 x 5mL).Coupling: A mixture of 7% DIPEA in DMF (3 ml) was added to a vial containing Fmoc-protected amino acid/HCTU (0.15mmol), and then transferred to the deprotected resin (NB: An additional 3mL of DMF was used to wash residual solution onto the resin). After sitting for 20 minutes with occasional stirring, the reaction vessel was drained and further washed with DMF (5 x 5mL).The synthesis cycle (deprotection with piperidine in DMF and coupling with a DIPEA/DMF/Fmoc-protected amino acid/HCTU solution) was repeated three more times with the appropriate Fmoc-protected amino acid. After the final step the resin was washed with methanol (2 x 5mL) and diethyl ether (2 x 5mL) and dried.Cleavage: TIPS (150 uL) and DMB (150 uL) were added to the resin followed by TFA (3 mL). After sitting for 30 minutes with occasional stirring, the mixture was filtered to isolate the resin. The filtrate was concentrated by a stream of nitrogen and then diethyl ether (40mL) was added to the filtrate. After being treated to vigorous shaking, the sample was isolated via centrifugation. (btw these documents are not created by me:))
PS The yield would be based on the mixture (of 2 major products in our sample, our pepitde and our peptide that has a AA missing presumably) otherwise there’ll masses for each after LC.. And then the purity will be determined from their relative ratios in the LC chromatogram.
