## 2 Chemical Synthesis of Nucleoside Di- and Triphosphates
### Early Methods for the Preparation of Nucleoside Di- and Triphosphates  
The first chemical synthesis of di- and triphosphates served mainly to determine and confirm the structure of nucleotides isolated from biological materials. The pioneering research in this field was carried out by Lord [PERSON], who, together with his collaborators, in the late 1940s and early 1950s, published a series of fundamental papers on the synthesis of an anhydride bond in nucleoside di- and triphosphates. Years of basic research have led to the development of new methods for the phosphorylation of alcohols and phenols with dibenzyl chlorophosphate (DBCP). It was found that this reagent could be used for both the phosphorylation of nucleosides and nucleotides (Scheme 2). Thus, when DBCP was allowed to react with a free \(5^{\prime}\)-OH group of the appropriately masked adenosine, dibenzyl adenosine \(5^{\prime}\)-monophosphate **1** was obtained. This compound could be partly deprotected by a careful treatment with diluted sulfuric acid in ethanol to form monobenzylated AMP **2**. The phosphorylation of this intermediate, again with DBCP, followed by hydrogenation, led to the preparation of the first synthetic nucleoside diphosphate, ADP (Scheme 2). It is noteworthy that the intermediate product **3** appeared to be a mixture of di- and tribenzyl diphosphates **3a** and **3b**, since one benzyl group was partly lost under the reaction conditions. Fortunately, there was no need for the separation of these products or a conversion of **3a** into **3b**, since the catalytic hydrogenation of this mixture furnished the desired adenosine \(5^{\prime}\)-diphosphate (ADP), which was isolated as an acridinium salt. After various modifications of this synthetic protocol, the product could be obtained in a 55% yield [39]. [PERSON] et al. [40; 41] also developed a method for the preparation of nucleoside 5\({}^{\prime}\)-triphosphates. The first synthesized compound was adenosine 5\({}^{\prime}\)-triphosphate (ATP), which helped confirm the assumed structure of a compound (tentatively identified as ATP) isolated from muscle extracts 15 years earlier. For this purpose, the aforementioned mixture of di- and triphenylzyl diphosphates **3a** and **3b** was used as the starting material. In the first step, fully protected diphosphate **3a** was converted into dibenzyl derivative **3b**. For selective monodebenzylation of the terminal phosphate residue (P\({}_{\beta}\)), a new reagent was developed, \(N\)-methylmorpholine [42]. After the short treatment (15 min) of the **3a**/**3b** mixture with this amine, Bn\({}_{2}\)ADP **3b** could be isolated from the reaction mixture. This was phosphorylated with DBCP to an electrically neutral tetrabenzyl ATP that was hydrogenated in order to remove the masking groups, and to furnish the final ATP as an acridinium salt with 37% yield. The relatively low yield of this method was accounted for by a rapid degradation of the anhydride bond in the completely masked 5\({}^{\prime}\)-triphosphate molecule, since adenosine 5\({}^{\prime}\)-diphosphate was observed as a by-product [40]. Another nucleotide, which structure was confirmed by chemical synthesis, was urine 5\({}^{\prime}\)-diphosphate (UDP). Initially, an attempt was made to use the same method as for ATP, but it proved to be ineffective as it resulted in large amounts of by-products, prob ably due to the more acidic nature of the uracil residue [43]. In this situation, [PERSON] et al. developed a new method for the formation of the P-O-P bond system in this particular compound, starting from benzyl uridine H-phosphonate diester **4** as a substrate (Scheme 3) [44] (preparation of H-phosphonate **4** from \(2^{\prime}3^{\prime}\)-\(O\)-isopropylideneuridine and dibenzyl pyro-H-phosphonate was described in an accompanying paper, also by [PERSON] et al. [45]).