Solid-phase peptide synthesis solvents

Since most aaAAs are hydrophobic in nature, peptides rich in aaAAs are generally restricted to study in organic solvents due to their low solubility in aqueous media. There have been very few examples of side-chain functionalized aaAAs that would allow for the synthesis of highly water-soluble peptides rich in aaAA content.3 This is primarily due to difficulty of synthesis, since side-chain functionalized derivatives must be orthogonally protected to allow for incorporation into solid-phase peptide synthesis. The harsh conditions, under which standard methods of aaAA synthesis are performed, make this a difficult task. 

The synthesis of some multiblock copolymers was attempted by successive polymerization using this iniferter technique. However, pure tri- or tetrablock copolymers free from homopolymers were not isolated by solvent extraction because no suitable solvent was found for the separation. In 1963, Merrifield reported a brilliant solid-phase peptide synthesis using a reagent attached to the polymer support. If a similar idea can be applied to the iniferter technique, pure block copolymer could be synthesized by radical polymerization. The DC group attached to a polystyrene gel (PSG) through a hydrolyzable ester spacer was prepared and used as a PSG photoiniferter (Eq. 53) [186] ... 

Cellulose is an unbranched polysaccharide consisting of 1,4-linked 3-D-glucose and can attain lengths of up to 15000 pyranose units. It neither dissolves nor swells in most solvents, but can be hydrolyzed upon prolonged treatment with acids. Cellulose powder was the first type of support to be used (with limited success) for solid-phase peptide synthesis [208]. Limitations were mainly due to the low loading (0.1 mmol/g) attained in these initial experiments, and the high reactivity of cellulose. Despite these... 

Standard solid-phase peptide synthesis requires the first (C-terminal) amino acid to be esterified with a polymeric alcohol. Partial racemization can occur during the esterification of N-protected amino acids with Wang resin or hydroxymethyl polystyrene [200,201]. /V-Fmoc amino acids are particularly problematic because the bases required to catalyze the acylation of alcohols can also lead to deprotection. A comparative study of various esterification methods for the attachment of Fmoc amino acids to Wang resin [202] showed that the highest loadings with minimal racemization can be achieved under Mitsunobu conditions or by activation with 2,6-dichloroben-zoyl chloride (Experimental Procedure 13.5). iV-Fmoc amino acid fluorides in the presence of DMAP also proved suitable for the racemization-free esterification of Wang resin (Entry 1, Table 13.13). The most extensive racemization was observed when DMF or THF was used as solvent, whereas little or no racemization occurred in toluene or DCM [203]. 

A difficult problem in utilizing enzymes as catalysts for reactions in a non-cellular environment is their instability. Most enzymes readily denature and become inactive on heating, exposure to air, or in organic solvents. An expensive catalyst that can be used only for one batch is not likely to be economical in an industrial process. Ideally, a catalyst, be it an enzyme or other, should be easily separable from the reaction mixtures and indefinitely reusable. A promising approach to the separation problem is to use the technique of enzyme immobilization. This means that the enzyme is modified by making it insoluble in the reaction medium. If the enzyme is insoluble and still able to manifest its catalytic activity, it can be separated from the reaction medium with minimum loss and reused. Immobilization can be achieved by linking the enzyme covalently to a polymer matrix in the same general manner as is used in solid-phase peptide synthesis (Section 25-7D). 

Based on the principle of the equal and simultaneous solvation of the polymer and the bound peptide chains in different solvents, Sheppard and coworkers developed a number of polyacrylamide-type supports for solid phase peptide synthesis 50 55). In this case, the crosslinked polymeric support, in addition to possessing the good mechanical characteristics like polystyrene, is much more structurally related to the peptide than in the case of polystyrene. The polar polyacrylamide support in this case is prepared by the emulsion copolymerization of a mixture of dimethylacrylamide (7), ethylenebisacrylamide (2) and acryloylsarcosine methylester (5), initiated by ammo-nium persulphatesl). 

Westall, E. C. and Robinson, A. B. (1970) Solvent modification in Merrifield solid-phase peptide synthesis. J. Org. Chem. 35, 2842-2844. 

Hyde, C. B., Johnson, T., and Sheppard, R. C. (1992) Internal aggregation during solid phase peptide synthesis. Dimethyl sulfoxide as a powerful dissociating solvent. J. Chem. Soc. Chem. Commun. 21, 1573-1575. 

This reagent, benzotriazol-l-yl-oxytripyrrolidinophosphonium hexafluorophosphate (14, PyBOP)P l (Scheme 4), was designed in order to avoid the formation of toxic HMPA during acylation. As with BOP, it is assumed that the first step is the carboxylic acid activation which involves formation of an acyloxyphosphonium salt.P This initial salt is then attacked by the benzotriazolyloxy anion to form the benzotriazolyl active ester which then reacts with the amino component. PyBOP can easily replace the BOP reagent and is especially suitable for solid-phase peptide synthesis. It is soluble in a wide range of solvents such as DMF, di-chloromethane, THF, and NMP. PyBOP is more useful in peptide synthesis on solid support than in solution. The byproduct, tris(pyrrolidino)phosphine oxide is partially water-soluble and is easily removed by washing. PyBOP is used under the same experimental conditions as BOP. Note that PyBOP is a white, crystalline and non-hygroscopic solid. It can be kept as a solid, but solutions of PyBOP cannot be stored for more than 24 hours. 

In batch SPPS (solid-phase peptide synthesis), the resin is contained in a vessel, usually equipped for bottom filtration. After a washing solvent or a reagent is added, the resin is mixed and the solvent or reagent is removed by filtration. Batch synthesizers use a variety of methods to mix the resin the wrist-action shaker, vortex mixing, nitrogen bubbling, or overhead stirring. 

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