CEM Liberty Blue User Guide

Cysteine
Cysteine is susceptible to epimerization at elevated temperatures in the presence of bases such as DIEA and NMM. If an
uronium activation strategy is being used, Cys should be coupled using the single 50 ° C 10 min coupling cycle with the
parameters listed in the table below.
Stage Temp (
o
C) Power (W) Time (sec) Delta T
1 25 0 120 2
2 50 50 480 1
Arginine
Arginine is susceptible to γ-lactam formation, greatly reducing the coupling efciency. For uronium activation, Arg is coupled
using an Arg double coupling cycle with the parameters listed in the table below.
First Coupling
Stage Temp (
o
C) Power (W) Time (sec) Delta T
1 25 0 1500 2
2 75 30 120 1
Second Coupling
Stage Temp (
o
C) Power (W) Time (sec) Delta T
1 75 30 300 2
Potential Side Reactions
Aspartimide Formation
Aspartimide formation is a common side reaction that occurs in peptides containing Asp followed by Asn, Asp, Gly, Thr,
or Ser. The addition of 0.1 M Oxyma Pure or 0.1 M HOBt to the deprotection solution will suppress the side reaction. When
using the UV monitoring option, Oxyma Pure and HOBt show signicant UV absorbance. When using acid-sensitive linkers
other than Cl-TCP(Cl), Oxyma Pure and HOBt can cause premature cleavage of the peptide. In these cases, the use of
Fmoc- Asp(OMpe)-OH in place of the more common Asp(OtBu) can reduce aspartimide formation without the addition of
Oxyma Pure/HOBt.
The Asp-Gly (DG) segment is particularly sensitive to base-catalyzed aspartimide formation. Simple addition of HOBt or
Oxyma Pure to the standard deprotection solutions may not be sufcient to prevent this side reaction. The use of less
aggressive deprotecting conditions or a Dmb protected dipeptide will prevent aspartimide formation from occurring.
Supression Strategy Microwave Method
5% (w/v) Piperazine w/ 0.1 M Oxyma Pure in EtOH:NMP (10:90) Two step room temperature deprotection:
1. Conventional initial deprotection
2. Conventional deprotection
Use Fmoc-Asp(OtBu)-(Dmb)Gly-OH dipeptide and any deprotection cocktail applicable for
standard peptides
Standard
Use Fmoc-Asp(OMpe)-OH and any deprotection cocktail applicable for standard peptides. Standard
Tetramethyl Guanidinium Capping
For extended coupling times (10 minutes or longer), tetramethylguanidinium capping of the free amine by HBTU (or other
uronium-type activators) is possible. This side reaction is detected as a truncation of target +101 on mass spec. Capping
by activator can be minimized by using a slight excess of other reagents (for example, 5 eq AA/4.5 eq HBTU/10 eq DIEA),
or by using alternate activator strategies (such as DIC/Oxyma Pure or DIC/HOBt).

CEM Liberty Blue Specifications

General

Brand	CEM
Model	Liberty Blue
Category	Laboratory Equipment
Language	English