Trityl-based linkers

Two trityl-based supports are presently in common use for the production of peptide adds by Fmoc SPPS 2-chlorotrityl chloride resin 7 (12) and 4-(chloro(diphenyl)methyl)benzoyl resin 8 (13). 

These resins are extremely moisture sensitive, and so it is essential that all reagents and glassware should be dried thoroughly before use. 2-Chlorotrityl chloride resin can be stored desiccated at room temperature 4-(chloro-(diphenyl)methyl)benzoyl resins should be generated from the precursor trityl alcohol immediately before use as described in Protocol 5. The chemical characteristics of 7 and 8 are identical. 

2-Chlorotrityl chloride resin is normally supphed with a displaceable chlorine content of 1.0-1.6 mmol/g. For the purposes of peptide synthesis, this substitution can sometimes be too high and can be reduced by treating the resin with a sub-stoichiometric amount of amino acid derivative and then capping imreacted sites with MeOH. 

Transfer resin as a damp cake to a round-bottomed flask. Add sufficient toluene to cover resin bed. 

Add AcCI (1 ml/g resin). Fit flask with refiux condenser and CaClj guard tube. 

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Trityl-based resins are highly acid-labile. The steric hindrance of the linker prevents diketopiperazine formation and the resins are recommended for Pro and Gly C-terminal peptides. Extremely mild acidolysis conditions enable the cleavage of protected peptide segments from the resin. These resins are commercially available as their chloride or alcohol precursors. The trityl chloride resin is extremely moisture-sensitive, so reagents and glassware should be carefully dried before use to avoid hydrolysis into the alcohol form. It is necessary to activate the trityl alcohol precursor and it is highly recommended to reactivate the chloride just before use see Note 4). After activation, attachment of the first residue occurs by reaction with the Fmoc amino acid derivative in the presence of a base. This reaction does not involve an activated species, so it is free from epimerization. Special precautions should be taken for Cys and His residues that are particularly sensitive to epimerization during activation (Table 2). 

A 9-phenylfluoren-9-yl-based linker can also be used for the attachment of alcohols. This Hnker shows improved acid stability compared with the trityl Hnker. The Hnker has been appHed to the synthesis of a peptide alcohol [71]. 

Cl Cl 6 p r= /= MeNHOH OH r= Y nA/om 0 0 [26] Only one example of nucleophilic cleavage with hydrox-ylamine has been described. Recently, many different hydroxylamine-based linkers have been described based on attachement of hydrox-ylamine to trityl-, Wang-, Sasrin- or Rink-resin. [26] [33] [175-177]... 

Of those in common usage, the 2-chlorotrityl 28 [74] and 4-carboxytrityl 29 [75] linkers give the least stabilized cations and are suitable for immobilization of carboxylic acids. They are ideal for Fmoc/tBu-based SPPS as their use avoids many of the side reactions that occur with standard benzyl-based linkers. Firstly, race-mization does not occur during loading of the resin with the C-terminal residue [79], as is the case with esterification to hydroxy-functionalized resins. Secondly, the bulky trityl cation does not cause alkylation side reactions with nucleophilic amino-acid side-chains. Thirdly, cysteine does not undergo racemization [80, 81]... 

For anchoring of the first amino acid onto hydroxymethyl-based resin, which upon cleavage provides a C-terminal carboxylic acids, it is recommended to use a protocol without the use of tertiary bases, such as DIEA. This type of protocol is designed to minimize the degree of self-acylation, hence, donble incorporation, and racemiza-tion of the first residne. The most common hydroxymethyl-based resins are Wang-type linkers (fcc Table 2). Two different protocols are described below for loading of hydroxymethyl-based resins (1) the symmetrical anhydride method and (2) the MSNT/Melm method. The MSNT/Melm method is recommended for difficnlt sitnations, which inclndes the attachment of amino adds that are prone to epimerization. For the synthesis of C-terminal adds where the first residue is either Cys or Pro, it is recommended to use the trityl-based resins. Many ofthe resins with a hydroxymethyl linker can be obtained with the first amino acid already preloaded. 

Only a few examples have been reported of the etherification of alcohols with resin-bound diarylmethyl alcohols (Entry 5, Table 3.30 Entry 5, Table 3.31 [564]). Diarylmethyl ethers do not seem to offer advantages over the more readily accessible trityl ethers, which are widely used as linkers for both phenols and aliphatic alcohols. Attachment of alcohols to trityl linkers is usually effected by treating trityl chloride resin or 2-chlorotrityl chloride resin with the alcohol in the presence of a base (phenols pyridine/THF, 50 °C [565] or DIPEA/DCM [566] aliphatic alcohols pyridine, 20-70 °C, 3 h-5 d [567-572] or collidine, Bu4NI, DCM, 20 °C, 65 h [81]). Aliphatic or aromatic alcohols can be attached as ethers to the same type of light-sensitive linker as used for carboxylic acids (Section 3.1.3). 

Trityl resins are particularly suitable for immobilization of nucleophilic substrates such as acids, alcohols, thiols, and amines. They are quite acid-sensitive and are cleavable even with acetic acid this is useful when acid-labile protecting groups are used. The stability of trityl resin can be tailored by use of substituted arene rings, as shown by chlorotrityl resin, which furnishes a more stable linker than the trityl resin itself. Steric hindrance also prohibits formation of diketopiperazines during the synthesis of peptides. Orthogonality toward allyl-based protective groups was demonstrated in the reverse solid-phase peptide synthesis of oligopeptides [30] (Scheme 6.1.4). 

Undoubtedly, the most important stereochemical feature is chirality. The [2]-rotaxane depicted schematically in Figure 2.12a is made with a macrocycle and a dumbbell that are not themselves chiral but oriented by appropriate substitution patterns on these components. Vogtle and co-workers described cycloenantiomeric rotaxanes that are based on the amide templating functionality (Figure 2.13).36 Racemic [2]-rotaxane 34 was synthesized in 20% yield by reaction of unsymmetrical acid dichloride 32 with trityl aniline 33 in the presence of macrocycle 31. The object and its mirror image are created by different sequences of the amide and sulfonamide linkers in both the dumbbell and the macrocyclic component. If the order of the atoms within the dumbbell is examined, in one enantiomer the sulfonamide nitrogen atom is... 

Coupling of 4-(4-hydroxymethyl-3-methoxyphenoxy)-butyric acid (HMPB, for synthesis of peptide acids) or p-[(R S)-a-[l- (9H- fluorenyl- methoxyform-amido]- 2,4- dimethoxybenzyl] - phenoxyacetic acid (modified Rink linker, for synthesis of carboxamide peptides) linkers to MBHA resin For Fmoc chemistry several types of solid supports are available, which include hydroxymethyl-based, aminomethyl-based, and trityl chloride resins. We describe the use of the MBHA resin. In this case the respective linker (to achieve peptide acid or amide) is coupled to the resin and first amino acid is then coupled to the linker. Attachment of the linker to the resin is a reaction between the carboxyl-group of the linker and amino-group of the MBHA resin. Commercially available resins with linkers already attached could also be used. 

A recently reported Hnker for alcohols is based on the 2-(diphenylmefhylsilyl)e-thoxymefhyl protecting group (64, Fig. 6). like other add-labile linkers, such as the THP or trityl Hnkers, it is stable under basic conditions [73]. The Hnker is also suitable for the attachment of amino functions. 

A new linker and a derived solid support based on the chlorofluorenyl residue li have been reported [82,83] that are analogous to the trityl supports in that carboxylic acids, amines, and phenols can be anchored. The resulting linkages proved to be more stable than those to the 2-chlorotrityl resins but could be cleaved by treatment with TFA. In the case of a carboxylic acid (Scheme 3), cleavage was achieved using 20% TFA in CH2Cl2-MeOH (9 1). 

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