Non-Polymeric Anchor-Assisted Synthesis

Among various liquid phase oligonucleotide synthesis (LPOS) methods, non-polymeric anchor-assisted approaches have emerged as innovative solutions for efficient oligonucleotide synthesis without the limitations posed by polymeric supports. This non-polymeric anchor system offers several key advantages. First, it enhances the solubility of the oligonucleotide intermediates, which facilitates efficient reaction kinetics during coupling cycles. Second, because the anchors are non-polymeric, the oligonucleotides can be recovered more easily through liquid-liquid extraction or phase separation, simplifying purification. Lastly, the use of non-polymeric anchors allows for the synthesis of longer and more complex oligonucleotide sequences compared to traditional polymeric-supported methods.

Common Synthesis Methods

• Ionic Liquid Tag-Assisted Synthesis

Soluble polymer supports face challenges such as low loading capacity and limited solubility when synthesizing longer sequences. To address these issues, non-polymeric supports can be used. For example, Donga et al.^[1] used ionic liquid-based supports to synthesize short oligonucleotides with different base compositions through phosphoramidite chemistry. These oligonucleotides achieved high yields and high purity, without requiring chromatographic purification before being cleaved from the support.

• Fluorous Tag-Assisted Synthesis

Fluorous tags are used to simplify the separation and purification of intermediates in the synthesis of biopolymers, as they allow the growing oligonucleotides to be easily purified through liquid-liquid or solid-liquid extraction procedures. It can be said that fluorous affinity purification is a very useful tool, especially for the separation of long oligonucleotides. Wada et al.^[2] synthesized dinucleoside phosphorothioates (oligonucleotide analogs) in a stereocontrolled manner (diastereoselectivity > 99:1) by introducing thymidine derivatives bearing a simple perfluoroalkyl tag (C₆F₁₃ or C₈F₁₇) at the 3'-end and diastereopure nucleoside 3'-O-oxazaphospholidine monomers. During the synthesis, the fluorous-tagged intermediates were easily purified by a simple fluorous solid-phase extraction process.

• Tetravalent Cluster Approach

Tetravalent cluster methods utilize both phosphoramidite chemistry and triester chemistry for synthesizing short oligonucleotides. During the synthesis process, the growing oligonucleotides, which are tetrahedrally branched from a pentaerythritol-derived core, are precipitated from methanol. This method has the advantage of good atom economy and is easy to analyze due to its small size and radially symmetric structure. Nevertheless, additional optimization is necessary to reduce side reactions and increase yield.

• Alkyl Chain-Assisted Synthesis

Alkyl chain soluble supports have also been used for liquid phase synthesis of biopolymers. For example, a research group^[3] has developed a high-yield method for synthesizing conjugated oligonucleotides that can easily couple reporter molecules with a wide range of polarities using an alkyl chain soluble support strategy. This method has the advantages of reducing cumbersome and convergent optimization of reaction conditions. Furthermore, all conjugated products, regardless of polarity, can be easily recovered and purified. These advantages significantly increase the efficiency of conjugated oligonucleotide synthesis.

As the demand for synthetic oligonucleotides continues to grow, non-polymeric anchor-assisted methods will play an increasingly important role in meeting the needs of the biotechnology and pharmaceutical industries. By providing flexible, scalable, and efficient alternative solid phase methods, they open up new possibilities for industrial-scale synthesis of complex oligonucleotide sequences. Alfa Chemistry is a leader in the field of non-polymeric anchor-assisted synthesis of oligonucleotides. If you have any questions or needs, please feel free to contact our technical team.

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