Of phosphoramidite

B. Studies of Equilibria and Reactions.—N.m.r. spectroscopy is being increasingly employed to study the mode and course of reactions. Thus n.m.r. has been used to unravel the mechanism of the reaction of phosphorus trichloride and ammonium chloride to give phosphazenes, and to follow the kinetics of alcoholysis of phosphoramidites. Its use in the study of the interaction of nucleotides and enzymes has obtained valuable information on binding sites and conformations and work on the line-widths of the P resonance has enabled the calculation of dissociation rate-constants and activation energies to be performed. 

The reaction of phosphoramidites (90) with the chlorophos-phoranes (91) produced the novel imidophosphates (92) containing both tetraco-ordinate and pentaco-ordinate phosphorus50. Phos-phorotropic migration to form (93) occurs for X = H or Me. 

The function of the acidic catalyst in alcoholysis reactions of phosphoramidites has not been elucidated yet, but low reactivity of... 

Tetrazolides of phosphorous acid esters or amides have been developed for the phos-phitilation of nucleosides. For instance, the tetrazolide of the diester of phosphorous acid (Af-tetrazolyldiethoxyphosphine) can be prepared either from diethylchlorophosphite and sodium tetrazolide (Method A) or from diethoxydiisopropylaminophosphine and two equivalents of tetrazole. The latter reaction (Method B) was undertaken to verify formation of a tetrazolide in the activation of phosphoramidites by tetrazole.[27]... 

The phosphitylation agent A-tetrazolyldiethoxyphosphine (phosphorous diester tetrazolide) can be made in situ from diethoxydiisopropylaminophosphine and two moles of tetrazole (tetrazole activation of phosphoramidites). 961... 

The use of click chemistry is a promising strategy as a postsynthetic ligation for nucleic acids in order to circumvent the time-consuming synthesis of phosphoramidites as DNA building blocks [31, 32]. This is particularly relevant for several fluorophores that are unstable under the acidic, oxidative, or basic conditions of automated DNA phosphoramidite chemistry and DNA workup. 

There is one more report on the synthesis of a library of phosphorus ligands on solid phase. Waldmann et al. prepared a library of phosphoramidites on beads (Fig. 36.5), but these were only applied in enantioselective C-C-bond formation. In fact, as two ligands need to be bound to the catalyst, the use of an immobilized monodentate ligands should most likely be avoided unless the proximity between the ligands is sufficiently close. In addition, crosslinking by the metal may have a negative impact on the permeability of the polymer for the substrate. 

Although addition of activated phosphoramidite to hemiacetals of manno-pyranoses under thermodynamic control has been reported to deliver exclusively a-phosphates in some cases,43 anomeric mixtures with preponderance of a-anomer have been reported in other examples.10,44 Since formation of phosphorotetrazolidite is a rate-limiting step of the process, initial activation of phosphoramidite followed by addition of nucleophilic hemiacetal should accelerate condensation and favour the formation of the thermodynamic a-product. Indeed, reaction of hemiacetal 101 with dibenzyl phosphorotetrazolidite assured exclusive a-selectivity of the resulting glycosyl phosphate 102.43 The accumulation in the reaction mixture of mildly acidic 1H-tetrazole, which is liberated upon reaction of tetrazolidite with hydroxylic component, could also favour predominant formation of the a-phosphate (Scheme 18, A). Conventional hydrogenolysis afforded the a-mannosyl phosphate 103. 

The X-ray structure of the Cut complex 21 of phosphoramidite 14 provides additional insight into a possible mechanism for stereocontrol (Fig. 7.3). The formation of the L2CuEt-enone complex involves substitution of the iodide in 21 for the alkyl moiety and of one of the ligands for the -coordinated enone. Coordination of RZnX results in the bimetallic intermediate 19 (Fig. 7.3). The absolute configuration of the two phosphoramidite ligands and the pseudo-C2-symmetric arrangement dictate the formation of (S)-3-ethyl-cyclohexanone. 

Reactions of allylic electrophiles with stabilized carbon nucleophiles were shown by Helmchen and coworkers to occur in the presence of iridium-phosphoramidite catalysts containing LI (Scheme 10) [66,69], but alkylations of linear allylic acetates with salts of dimethylmalonate occurred with variable yield, branched-to-linear selectivity, and enantioselectivity. Although selectivities were improved by the addition of lithium chloride, enantioselectivities still ranged from 82-94%, and branched selectivities from 55-91%. Reactions catalyzed by complexes of phosphoramidite ligands derived from primary amines resulted in the formation of alkylation products with higher branched-to-linear ratios but lower enantioselectivities. These selectivities were improved by the development of metalacyclic iridium catalysts discussed in the next section and salt-free reaction conditions described later in this chapter. 

Concurrent with studies on cyclometalation, studies on the effects of the structure of phosphoramidite ligand had been conducted. Several groups studied the effect of the stmcmre of ligand on the rate and selectivity of these iridium-catalyzed allylic substitutions. LI contains three separate chiral components - the two phenethyl moieties on the amine as well as the axially chiral BINOL backbone. These portions of the catalyst structure can control reaction rates by affecting the rate of cyclometalation, by inhibiting catalyst decomposition, or by forming a complex that reacts faster in the mmover-limiting step(s) of the catalytic cycle. 

For the copper-catalyzed 1,4-addition to 2-cyclohexen-l-one, other alkylmetal reagents have also been employed, achieving high enantioselectivity in some cases [Eqs. (3.2)-(3.4)]. Recently, one example appeared that utilized diphenylzinc as the nucleophile in the presence of phosphoramidite ligand 1 to produce highly enantio-enriched 3-phenylcyclohexanone [Eq. (3.5) 94% ee]. ... 

Figure 4.5 Scheme for stepwise synthesis of oligonucleotides via coupling of phosphoramidite-functionalized nucleosides. Reprinted from Noro et al. (2005). Copyright 2005 American Chemical Society. 

The copper-catalyzed enantioselective Michael addition of organometallic reagents to enones was the first successful application of phosphoramidite chiral ligands in catalysis [4, 43]. Since this early report, substantial enhancement of the enantioselectivity and/or of the substrate scope has been achieved through an untiring effort to optimize the ligand structure [5a, 44]. 

It was later found that dialkyl(dialkylamino) phosphites (phosphoramidites, (RO)2PNR2 [114]), which are stable towards air and water and can be stored for longer, can readily be converted into the phosphinic chlorides (RO)2PCl by treatment with dimethylaniline hydrochloride, or into the corresponding tetrazolides by treatment with tetrazole. Tetrazolides (RO)2P-(l-tetrazolyl) had proven excellent reagents for the phosphorylation of nucleosides [113], and the treatment of phosphoramidites with alcohols in the presence of tetrazole was found to be a satisfactory method for the rapid preparation of trialkyl phosphites [115,116]. 

The rhodium-catalysed enantioselective 1,4-addition of arylboronic acids to the bifunctional Michael acceptors (192) in the presence of phosphoramidites (194) occurs regioselectively at the endocyclic C=C bond and in up to 95% ee. The presence of KOH is required to increase the reactivity so that less boronic acid and lower reaction temperatures can be used.241... 

Huttenloch and co-workers prepared an entire library of phosphoramidite ligands on solid phase.50 However, these were only used for conjugate addition reactions. 

Alternatively to the DNA modifications in the previous two sections where the chromophore was attached to one of the four DNA bases, chromophores can be incorporated as artificial DNA bases substituting a natural base or even a whole base-pair. There is a large number of recently reported syntheses of chromophores as DNA base surrogates, e.g. flavine derivatives [26] and thiazole orange derivatives [42]. Additionally, a variety of phosphoramidites as DNA building blocks for the introduction of fluorophores into DNA are commercially available, e.g. acridine derivatives. Clearly, the synthetic protocols for this kind of DNA modification do not follow a principle strategy which can be applied in a versatile fashion, as is the case for the DNA base modifications mentioned in the previous sections. It is important to point out that in many cases it turned out to be useful to replace the 2 -deoxyribose moiety with acyclic linker systems. This was also the case during our attempts to synthesize ethidium-modified DNA, which will be described here briefly. 

Over the last 20 years methods for the chemical synthesis of oligodeoxyribonucleotides and oligoribonucleotides have become available. The use of phosphoramidite chemistry, the introduction of solid-phase synthesis techniques and the possibility of automated synthe.sis has proved to be a powerful stimulus to streamline and simplify chemical procedures.23 ... 

Kool described the synthesis of the phosphoramidite derivative of a C-nucleoside incorporating a porphyrin moiety, (146)." His approach was to assemble the porphyrin de novo on the sugar moiety starting from 3,5-bis-O-toluoyl-protected deoxyribose-Cl-carboxaldehyde, benzaldehyde and dip-yrromethane under Lindsey conditions. Similarly, to fluorescently label oligonucleotides, Burgess reported the synthesis of phosphoramidite (147), for which the nucleoside precursor was prepared from 5-ethynyl thymidine and iodofluorescein via Sonogashira s coupling procedure. ... 

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