Facile synthesis of novel mutual derivatives of nucleosides and pyrimidines by regioselectively chemo-enzymatic protocol

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Abstract

We established a facile regioselectively chemo-enzymatic synthesis procedure for the preparation of mutual derivatives of nucleosides and pyrimidines by sequential Markovnikov addition and acylation. Firstly, pyrimidine derivatives containing vinyl ester group were synthesized from pyrimidines and divinyl esters through Markovnikov addition catalyzed by K2CO3 in DMSO at 80 °C, and the yields were ranged from 50% to 87%. Then regioselective acylation of ribavirin and cytarabine with pyrimidine vinyl ester was catalyzed by CAL-B (immobilized lipase from Candida antarctica) in anhydrous acetone. Reaction conditions of enzymatic acylation including enzyme resource and solvents were optimized. A series of mutual derivatives of nucleosides and pyrimidines were synthesized successfully and characterized with NMR, IR, and HRMS. This chemo-enzymatic protocol involving sequential Markovnikov addition and acylation provided a novel way of synthesizing complicated functional compounds regioselectively which was hard to be achieved either by chemical or by enzymatic methods.

Graphical abstract

A facile regioselectively chemo-enzymatic synthesis procedure for the preparation of novel mutual derivatives of nucleosides and pyrimidines was developed by sequential Markovnikov addition and acylation.

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Introduction

Nucleosides are the most frequently used effective class of antiviral agents, with over twenty drugs currently approved for the treatment of viral diseases and a number of candidates in the clinical trials.1, 2 Consequently, the intense search for new nucleoside derivatives attracted extensive attention. For example, Mackman synthesized a novel nucleoside phosphonate which had anti-HIV activity.3 Being bioactive molecules, pyrimidines are important components of the biological macromolecules, such as DNA and RNA. So introducing the pyrimidines into nucleoside derivatives may result in the discovery of a number of novel derivatives with potential antitumor and antiviral activities. The explosion of new approaches for their synthesis and most importantly, their selective synthesis is an interesting subject of organic and bioorganic chemistry.

Enzyme-catalyzed reactions are widely recognized as superior to conventional chemical methods in selective modification of polyfunctional substrates owing to mild reaction conditions, high catalytic efficiency, inherent selectivity, and simple downstream processing. Therefore, many research groups have paid much effort in the area of enzymatic synthesis of nucleoside derivatives.4, 5, 6, 7, 8, 9 Transesterification is the most common method to synthesize nucleoside derivatives. For instance, Hanson regioselectively synthesized the prodrug of Lobucavir by screening the solvents and enzymes.10

Many of the researches in this field, however, are focused on acylation reaction type which hardly provides more procedures for preparing complicated functional compounds. Therefore, the combination of the enzymatic reaction and diversified chemical reaction will be a smart choice. Gotor chemo-enzymaticly synthesized 3′- and 5′-carbazyl nucleoside derivatives.11 Tanaka also synthesized antiviral carbocyclic nucleosides using rhizopus delemar lipase.12 Addition reaction is one of the most basic types of reactions in organic synthesis. Exhilaratingly, we have found that some enzymes are able to catalyze general addition reactions such as Michael addition and Markovnikov addition.13 It provided an easy way to introduce various functional groups into N-heterocycles including pyrimidine or purine. Our group has demonstrated a facile method to synthesis complex compounds containing the sugar moiety and heterocycles using the combination of enzyme-catalyzed addition and acylation reactions.14

Here, we expected to regioselectively synthesize novel and complicated nucleoside derivatives containing bioactive pyrimidines. Two nucleosides including ribavirin and cytarabine, and three pyrimidines are selected as the starting substrates for the mutual derivatives of nucleoside and pyrimidines. We found the step of Markovnikov addition would carry out very well while using chemical catalyst such as K2CO3. Furthermore, reaction conditions for the base-catalyzed addition and enzymatic selective acylation were optimized, respectively. As a result, twelve mutual derivatives of nucleoside (ribavirin or cytarabine) and pyrimidines were regioselectively prepared in good or moderate yields using the chemo-enzymatic strategy combining Markovnikov addition and acylation.

Section snippets

Influence of catalysts on Markovnikov addition

Markovnikov addition is one of the most basic types of reactions in organic synthesis which can facilely introduce various functional groups into N-heterocycles. Divinyl dicarboxylates are selected for the preparation of pyrimidines derivatives which will be subjected to the second-step transformations (Scheme 1). Markovnikov addition between thymidine and divinyl adipate was chosen as model reaction. Three kinds of bases were screened in order to find the most efficient one for the preparation

Conclusion

By optimizing the conditions of sequential two-step reactions, a new synthetic strategy of mutual derivatives bearing nucleosides and pyrimidine bioactive moieties via chemical Markovnikov addition and enzymatic acylation was established. Our results clearly demonstrated that the reaction between pyrimidine and vinyl ester could be efficiently promoted by 10 mol % K2CO3 in DMSO at 80 °C, and the further transesterification between ribavirin or cytarabine and fatty acid pyrimidine vinyl esters was

Materials

Lipozyme® (E.C. 3.1.1.1, an immobilized preparation of lipase from Mucor miehei, 42 μ/g), lipase from porcine pancreas (PPL) (E.C. 3.1.1.3, Type II, powder, 30–90 μ/mg), lipase from C. cylindracea (CCL) (E.C. 3.1.1.3, powder, 2.8 μ/mg) was purchased from Fluka. C. antarctica lipase acrylic resin (CAL-B) (E.C. 3.1.1.3, an immobilized preparation of lipase from C. antarctica on macroporous acrylic resin, 10,000 μ/g,) and lipase Type VII from Candida rugosa (CRL) (E.C. 3.1.1.3, powder, 706 μ/mg) were

Acknowledgments

The financial support from the National Natural Science Foundation of China (Nos. 20572099, 20704037), Ph.D. Programs Foundation of Ministry of Education of China (20060335031), and the Zhejiang Provincial Natural Science Foundation (Project No. 2008-Y407086) is gratefully acknowledged.

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