Novel methodologies for solid and liquid phase peptide synthesis
- Autores: Vida de la Paz del Mar Castro Pinzón
- Directores de la Tesis: Fernando Albericio Palomera (dir. tes.), Hortensia Rodríguez Cabrera (codir. tes.)
- Lectura: En la Universitat de Barcelona ( España ) en 2014
- Idioma: español
- Tribunal Calificador de la Tesis: Pedro Romea Garcia (presid.), Miriam Royo Expósito (secret.), María Margarita Suárez Navarro (voc.)
- Materias:
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- Resumen
Peptides play a decisive role in many physiological processes, whether as neurotransmitters, hormones or antibiotics, but also exhibit a broad range of biological activities as antimicrobial, antioxidative, antihypertensive, cytomodulatory and immunomodulatory agents in a living body system. With the needed research and development, there exists enormous opportunities to effectively harness these diverse functionalities of bioactive synthetic peptides for the treatment, prevention and mitigation of different medical conditions.
In 1963, Merrifield introduced the concept of SPPS reporting the first efficient production of a tetrapeptide using polystyrene (PS) resin as a solid matrix where the peptide chain is grown by being covalently attached at one end to the functionalized support. Thanks to Merrifield¿s pioneering work, nowadays peptides have a fully established method of chemical synthesis based on solid-phase techniques in combination with progress in e¿ective protection and deprotection methods. These techniques have combined well with automated and combinatorial strategies to comprise a substantial fraction of current peptide synthesis in both the academic and industrial ¿elds.
Some factors related to SPPS methodology have contributed to the popularity of the technique: (i) The ease of the chemistry involved, reactions can be accomplished in just three steps: addition of reagents, filtering, and washing the resin, thus allowing many simple automated procedures to be developed; (ii) The elimination of purification steps en route, for each step of a multiple-step synthesis, the only purification needed is a resin-washing step. Only the final cleaved product needs to be purified; (iii) In a solid-phase synthesis, high concentrations of reagents can be used to drive reactions to completion; and (iv) The straightforward nature of parallel solid-phase synthesis.
Despite the undeniable success of this synthetic methodology, it still presents certain drawbacks. The most notable drawback is the heterogeneous reactions conditions, which can exhibit several of the following problems: non-linear kinetic behavior, unequal distribution and/or access to the chemical reaction, solvation problems, the use of insoluble reagents or catalysts, pure synthetic problems associated with solid phase synthesis. In addition, the range of chemistry available on solid phase is clearly limited and it is difficult to monitor the progress of the reaction when the substrate and product are attached to the solid phase. Because of these solid phase synthesis limitations, alternative methodologies have been reported.
Alternative approaches consisting of soluble polymer-tagged liquid-phase reactions have also been developed to facilitate reaction work-up and product isolation, with some success in preparing bioactive peptides. The liquid phase synthesis concept was introduced by Mutter in 1972 as a hybrid approach between solution and solid phase methods. The central feature of this methodology is that it combines the advantages that classic organic synthesis in solution offers with those that solid-phase synthesis can provide, through the application of a soluble polymer. In principle, liquid-phase methods have no scale limitations, and the reactions can be monitored directly using standard analytical techniques. In contrast, the major drawbacks on the LPPS global process are the purification methods to remove the remaining reagents; hence, SPPS has developed strikingly fast since then.
Taking into account all the aforementioned, it can be concluded that both SPPS and LPPS have scopes and limitations; therefore, further research work in the development and/or improvement of synthetic methodologies is still necessary. In work herein presented, some improvements for SPPS have been developed, Moreover, an approach to a recently created new technology platform based on membrane assisted-LPPS has also been established.
Supported peptide synthesis is based on the polymer, the cleavable linker, the protection scheme and the coupling reagents. The thesis presented herein focuses on this type of synthesis: to develop a safe linker for a free side-reactions preparation of peptides The main specific objectives are; to carry out a systematic study to establish a standard, efficient and mild protocol to prepare azide-solid supports, trying to assess the scope and limitations of imidazole-1-sulfonyl azide (ISA¿HCl) as a diazo-transfer reagent in several resins. To introduce a new concept of linker. In this case, the linker is anchored to the solid support through a stable triazole moiety via Cu(I) catalyzed azide/alkyne cycloaddition (CuAAC) (Click Chemistry reaction) and with methoxy groups as the only activating groups of the phenyl ring. To compare the efficiency of the new resin in solid phase synthesis (SPS) with that of Wang resin.
