Interpretación según perspectiva semiolingüística

phenyl protecting groups have led to the use of other phosphorylating reagents.129 By using tetrabenzylpyrophosphate and n-butyllithium, Frost et al.129 avoid using the unstable dibenzyl phosphorochloridate reagent and could deprotect the dibenzyl phosphate carbohydrate via hydrogenolysis using palladium on carbon.

Other methods involving the pyrophosphate moiety include adding the disubstituted phosphorochloridate to a solution of tributylammonium hydrobenzoin cyclic phosphate (THCP), and tributylamine as shown by Aimi et al.130 The desired carbohydrate is then added to the reactive pyrophosphate moiety.131 This method, however, suffers from moderate yields and poor atom economy.

Phosphate production is not only limited to the reaction of electrophilic reagents with a nucleophilic hydroxyl group. Hilton et al.132 developed a method where potassium diethyl phosphite in liquid ammonia has been shown to substitute alkyl triflates and form phosphate esters (scheme 2.05).132 The phosphate ester product represents a formal oxidation of phosphorus.

Scheme 2.05. Example of electrophilic phosphorylation.132 H OTf H OPO(OEt)2 K O P(OEt)2 NH₃ 85%

Reports of phosphorylating reagents using phosphorus(III) have become frequent, mainly due to Beaucage and Caruthers133 _{developing methods involving phosphoramidite in the}

early 1980s. In a recent example by Graham and Pope (scheme 2.06),117 the C5-hydroxyl of D-riboside 2.04 attacks the reactive tetrazole and phosphoramidite producing a phosphite triester, which is subsequently oxidised by tert-butyl hydroperoxide (TBHP) to the phosphate ester 2.05. If recovery of starting D-riboside 2.04 is allowed for, then the yields improve further, exceeding 85%.117 A main point of difference between the

phosphoramidite and most phosphorus(III) phosphorylating methods is the removal of the need to rely on alkaline conditions to catalyse the reaction.

Scheme 2.06. Phosphorylation of C5-hydroxyl using the phosphoramidite

method.117 O O HO OH OH N N N HN R R= p-PhNO₂ BnO P BnO N(i-Pr)₂ i) MeCN ii) TBHP CH2Cl2 O O HO OH (BnO)₂OPO 60% 2.04 2.05

Variations of the phosphoramidite method include changing the oxidant to m- chloroperoxybenzoic acid (m-CPBA) and the phosphate protecting groups to ethyl or

tert-butyl.134,135

2.1.4. Literature deprotection of phosphate esters:

As mentioned, the diphenyl and dibenzyl phosphate ester protecting groups can be removed by hydrogenolysis with H2 with platinum and palladium on carbon

respectively.124,125 Dimethyl and diethyl phosphate ester protecting groups can be cleaved using trimethylsilyl bromide (TMSBr) and NEt3 in CH3CN.136 Similar deprotections of

alkyl phosphate esters have been reported by employing NaI in acetone137 or the generation of Me3SiI in situ from Me3SiCl and NaI in CH3CN.138-141

One of the problems we could foresee with the alkyl deprotection step is that the phosphate ester moiety could also be cleaved at the carbohydrate moiety. In terms of reactivity we believe this should follow cleavage of the phosphate ester alkyl groups. Taking care in ending the reaction before the phosphate ester has time to cleave is of up- most importance. Also there is a possibility, when using NaI in acetone, that after the first

alkyl group cleaves the resultant sodium salt may precipitate, thereby inhibiting the cleavage of the second ethyl group and yielding only the mono-protected rCdRP product 2.06 (figure 2.01). This has been discussed in literature previously.137

2.06 O OH HO HO H N P O- O O O -_O

Figure 2.01. Mono-protected product.

2.2. Phosphorylation of

ribonolactone:

Following the retrosynthesis plan shown in scheme 2.02, route B, we went about attempting to synthesise D-ribonolactone-5-diphenyl phosphate 2.07. With two

unprotected secondary hydroxyl groups exposed, selectively phosphorylating the primary hydroxyl group of D-ribonolactone 2.03 was paramount. Diphenyl phosphorochloridate was selected for this task due to its sterically bulky phenyl groups. In a procedure based on work done by Tener and Khorana,122 D-ribonolactone 2.03, imidazole and diphenyl phosphorochloridate were stirred in dry CH2Cl2 for several hours at room temperature.

Thin layer chromatography (TLC) monitoring by spotting crude aliquots of reaction mixtures on silica plates proved difficult due to the poor solubility of 2.03 in CH2Cl2. The

polar nature of 2.03 demanded a solvent or conditions that would improve solubility and allow reactivity with diphenyl phosphorochloridate. Keeping the other reaction condition parameters constant, the solvent was altered from CH2Cl2 to the more polar N,N-

dimethylformamide (DMF). However, these conditions failed to produce the phosphorylated product 2.07. A low Rf spot on TLC, however, was apparent unlike the

reaction carried out in CH2Cl2. Upon work-up, this spot disappeared when the crude

reaction mixture was washed with aqueous HCl. Concentrating the aqueous layer produced a cream coloured syrup, which was later determined to be the unreacted starting material 2.03 by 1H NMR (D2O). The organic phase of the work-up contained diphenyl

shift change from –4.1 ppm to –9.9 ppm was noted with the hydrolysis of diphenyl phosphorochloridate.

Changing the solvent to tetrahydrofuran (THF) did not produce phosphorylated product 2.07. Neither did warming the reaction mixture to 40 °C over a period of several hours with or without the use of an ultrasonic bath. Increasing the ratio of phosphorylating reagent and imidazole or use of additional base in the form of pyridine also did not produce any phosphorylated product 2.07. Finally after refluxing with a high ratio of phosphorylating reagent and imidazole to D-ribonolactone 2.03, in ultrasonic conditions

(scheme 2.07), a decision was made to explore other avenues to a phosphorylated precursor (scheme 2.02), since no product (N. P.) was formed.

It is likely the solvents used did not fully solubilise D-ribonolactone 2.03, or produce

conditions for reactivity with the phosphorylating reagent. This is evident by the crude yield of recovered starting material, D-ribonolactone 2.03 (~90 %, scheme 2.07). Approximate yield of diphenyl phosphate 2.08 (70%) was based on the number of moles