Use of Hydroxylamines, Hydroxamic Acids, Oximes and Amines as Nucleophiles in the Zbiral Oxidative Deamination of N-Acetyl Neuraminic Acid. Isolation and Characterization of Novel Mono- and Disubstitution Products

Abstract

The oxidative deamination of N-nitroso N-acetylneuraminic acid (NeuAc) derivatives is a useful reaction for the formation of 5-desamino-5-hydroxy NeuAc derivatives and their stereoisomers. We demonstrated previously that replacement of the classical nucleophile in these reactions, acetic acid, by phenols resulted in a novel double displacement process with substitution of the acetoxy group at the 4-position taking place in addition to that of the 5-acetamido group, for which we postulated a mechanism centered on the formation of a highly reactive vinyl diazonium ion. We now extend these studies to encompass the use of hydroxylamine-based systems and weakly basic amines as nucleophile. We find that the nature of the product depends significantly on the pKa of the nucleophile, with the more acidic species typically affording only substitution at the 5-position, while the less acidic species give mixtures of elimination products and disubstitution products. The use of aniline as nucleophile is of particular note as it affords a novel aziridine spanning positions 4- and 5- of the neuraminic acid skeleton.

Graphical Abstract

Introduction

The oxidative deamination of N-acetyl neuraminic acid (NeuAc) glycosides 1 by treatment of the corresponding N-nitrosoamides 2 with sodium trifluoroethoxide followed by the addition of acetic acid resulting in the replacement of the C-N bond by a C-O bond with retention of configuration is a practical and convenient synthesis of 3-deoxy-D-glycero-D-galactononulosonic acid (KDN) glycosides 3 (Scheme 1, X = OAc) first described by Schreiner and Zbiral,¹ and extended to encompass NeuAc thioglycosides in our laboratory.² The overall transformation of 2 to 3 also may be achieved by simple thermolysis, also with retention of configuration, but such conditions with their seemingly narrower scope have been less widely explored.³

Scheme 1.

Oxidative Deamination of NeuAc

As originally described by Schreiner and Zbiral¹ the only nucleophiles employed were acetic acid (Scheme 1) and hydrogen azide, with the latter affording 5-azido-5-desacetamido-NeuAc derivatives, also with retention of configuration. In our laboratory we extended the range of nucleophiles to include alternate carboxylic acids (specifically levulinic acid),⁴ hydrogen fluoride,⁵ trifluoromethanesulfonic acid,⁶ thioacetic acid,⁵ thiophenols,⁷ and provided the second step is conducted in the presence of fluoroboric acid, alcohols,⁵ with each substitution taking place with retention of configuration (Scheme 2). In a series of experiments designed to probe mechanism of substitution with retention of configuration we conclusively eliminated stereodirecting participation by esters at the 4- or 7-positions from consideration and hypothesized that participation by the pyranosidic ring oxygen via an intermediate 1-oxabicyclo[3.1.0]oxahexanium ion underlies the stereochemical outcome of these reactions.⁴ Seeking to further expand the range of nucleophiles compatible with the oxidative deamination we recently explored the use of phenols and uncovered a new reaction manifold with concomitant substitution at the 4-position. Thus, replacement of acetic acid as nucleophile by phenol under our otherwise standard conditions resulted in the isolation of the 4,5-disubstituted derivative 4 as major product with retention and inversion of configuration at the 4- and 5-positions, respectively. Conversely, the use of β-naphthol as nucleophile gave the crystallographically-established tricyclic system 5 as major product with inversion and retention of configuration at the 4- and 5-positions, respectively (Scheme 2).⁷

Scheme 2.

Nucleophiles Employed Previously in the Deamination of NeuAc

We understand this ensemble of results in terms of the initial formation of a diazonium ion 6 on treatment of 2 with trifluoroethoxide that is in a pH dependent equilibrium with the diazoalkane 7 (Scheme 3). The intermediacy of the diazonium ion 6 is established by the isolation of a typical azo dye as byproduct (not shown) in the reaction with β-naphthol,⁷ whereas the equilibrium 6 and 7 rests on the isolation of 5α-deuterio 3 (not shown) when deuterioacetic acid is used as nucleophile.⁴ Addition of nucleophiles with pKa <~8 results in the loss of nitrogen with formation of the 1-oxabicyclo[3.1.0]hexanium ion 8, and overall substitution with retention of configuration as in 9, ie, the classical reaction described by Schreiner and Zbiral¹ and widely employed by ourselves and others as a means of entry into KDN and other sialosides.^{2, 5, 8–10} With the less acidic phenols as nucleophiles the equilibrium favors the diazoalkane 7 resulting in the elimination of acetate from the 4-position giving an alkenediazonium ion 10. Kinetic attack on the β-face of this alkenediazonium ion, perhaps via the ²H_O half-chair conformer, gives an adduct 11 which following protonation and displacement of nitrogen affords the final products. The kinetic mode of reaction of 9 is exemplified by the use of β-naphthol as nucleophile resulting ultimately in the isolation of 5. Thermodynamic attack occurs on the α-face of the alkenediazonium ion 10, perhaps via the ^OH₂ half-chair conformer, to give an adduct 12 from which disubstituted products such as 4, observed with phenol as nucleophile, flow (Scheme 3).⁷

Scheme 3.

Classical and New Reaction Manifolds for the Deamination of NeuAc Dependent on the pKa of the Added Nucleophile

We now report on the on the use of further nucleophiles of varying pKa in the deamination of NeuAc with a view to providing further insight into the overall mechanistic scheme and increasing the variety of NeuAc-like structures available for medicinal chemistry purposes.

Results

The first nucleophile studied in this investigation, in view of its well-known and widely exploited nucleophilicity, was 1-hydroxybenzotriazole (HOBt). A single major product 13 was isolated in 48% yield that arose from substitution of the nitrosoacetamide function with retention of configuration (Table 1, entry 1). A mixture of the two elimination products 14 and 15 was also formed in this reaction, as determined by mass spectrometry and NMR spectroscopy of the crude reaction mixture, albeit they were not isolated and quantified owing to the relative complexity of the reaction mixture, which is typical of the general reaction class. Turning to the use of N-hydroxyphthalimide as nucleophile, we isolated 36% of the standard Zbiral-type product 16, and 32% of the disubstitution product 17. Again, significant amounts of the elimination products 14 and 15 were formed in this reaction (Table 1, entry 2). With the less acidic acetohydroxamic acid¹¹ as nucleophile a bicyclic product 18 was formed in 21% yield alongside a single elimination product 15, which was isolated in 28% yield (Table 1, entry 3). Use of N-(p-methoxybenzyl)acetohydroxamic acid as nucleophile on the other hand gave only 18% of the combined elimination products 14 and 15, and 21% of the 4-deoxy-5-desacetamido non-4-eneulosonic acid derivative 19 (Table 1, entry 4), a class of sialic acid derivatives only previously accessed by Takahashi and coworkers by degradation of the NeuAc framework to a 7-carbon synthon, followed by reassembly.¹² With the more acidic N-hydroxyimide, N-Boc acetohydroxamic acid, the elimination products 14 and 15 were the predominant products isolated in a combined yield of 41%, while 19 was isolated as a minor product in 10% yield. An inseparable mixture of the ketone 21 and the enone 22 in 17% yield Table 1, entry 5) was also isolated. Turning to the use of oximes as nucleophiles, application of p-nitrophenylbenzaldehyde oxime afforded the adduct 20 in 26% yield along with a mixture of elimination products 14 and 15 in 24% yield, together with the inseparable mixture of the ketone 21 and the enone 22 in 8% yield Table 1, entry 6). With the less acidic and more hindered acetone oxime on the other hand only a mixture of the elimination products 14 and 15 could be isolated from the reaction mixture in 38% yield (Table 1, entry 7).

Table 1.

Application of Hydroxylamine Derivatives and Amines as Nucleophiles in the Zbiral Reaction^a

^a)Unless otherwise stated all reactions were conducted at −10 °C in dichloromethane. b) Nucleophiles were employed in 10–20 fold excess as detailed in the Supporting Information. c) The elimination products 14 and 15 are the major side products in all reactions as determined by inspection of the crude reaction mixtures by mass spectrometry and NMR spectroscopy, albeit they were not isolated and quantified in every case. .

In addition to the use of hydroxylamine derivatives as nucleophiles we also briefly investigated that of the nitrogen nucleophiles aniline, indole, and 3,5-dimethylpyrazole. With aniline a complex reaction mixture was observed from which we isolated the aziridine 23 in 49% yield (Table 1, entry 8), while the use of indole afforded only the elimination products 14 and 15, albeit with an excellent yield of 80% (Table 1, entry 9). 3,5-Dimethylpyrazole was not sufficiently soluble in dichloromethane for use as nucleophile, but was soluble in hexafluoroisopropanol, a weakly nucleophilic solvent^13–15 we had previously employed to good effect for the addition of the poorly soluble 6-hydroxyquinoline to the vinyldiazonium ion 20 and resulting in the isolation of a heterocyclic analog of tricyclic product 5 in good yield.⁷ Unfortunately, addition of 3,5-dimethylpyrazole in hexafluoroisopropanol to a typical dichloromethane solution of the N-nitrosoamide 2 gave a complex reaction mixture from which none of the anticipated products could be recovered. We did, however, isolate from this mixture a product 24 resulting from the combination of two molecules of hexafluoroisopropanol with the activated nitrosoamide, albeit in only 9% yield (Table 1, entry 10).

Structural Elucidation

The structures of elimination products 14 and 15 was confirmed by comparison of their spectra with those of authentic samples,⁷ while the structures of adducts 16, 17, and 19 follow without complication from inspection of their NMR spectroscopic and mass spectrometric data. The structure of the adduct 20 is assigned by comparison of its spectral data with that of 15 and an analogous product isolated from the application of 4-nitrophenol as nucleophile.⁷ The structure of enone 22 was assigned by comparison with an authentic sample,⁷ while that of ketone 21 follows from its gradual decomposition into 22, which also prevented it from being isolated in pure form.

The configuration of the N-hydroxybenzotriazole adduct 13 is readily apparent from its ¹H NMR spectra, which reveal the all-equatorial nature of the substituents at the 4-,5-, and 6-positions around the pyranoside ring. That 13 resulted from the attack of the ambident nucleophile N-hydroxybenzotriazole on oxygen and not nitrogen – resulting in the formation of betaine 25 – was ascertained by treatment with zinc in acetic acid followed by acetylation to give the known KDN derivative 26 in 97% overall yield (Scheme 4).

Scheme 4.

Confirmation of the Structure of the N-Hydroxybenzotriazole Adduct 13.

The structure of 18 is based on the HMBC correlation of the imidate carbonyl C with H5 and not with H4, which excludes the regioisomer 27 (Figure 1). The cis-fused ring junction and chair-like conformation of the pyranose ring in 18 is apparent from inspection of its NMR spectroscopic data including analysis of the complete set of ³J proton-proton couplings in the H3a,e-H6 spin system. Finally, the fusion of the heterocylic ring to the β-face of the pyranose is confirmed by nOe correlations between H5 and H’s 3 and 7 on the α-face of the molecule (Figure 1). Aziridine 23 was assigned on the basis of the nOe correlations between the side chain proton H7 and the ring protons H4 and H5, which place the aziridine ring on the opposite face of the pyranoside ring to the side chain. The exoconfiguration is clear from the nOe correlations between the ortho-protons on the phenyl ring and H’s 4 and 5 (Figure 2).

Figure 1.

HMBC and Nuclear Overhauser Correlations Supporting the Structure of 18 as Compared to the Regioisomer 27.

Figure 2.

Nuclear Overhauser Correlations Establishing the Structure of 23.

Finally, the configuration and conformation of the hexafluoroisopropanol adduct 24 are established by analysis of NMR coupling constants and nOe correlations. Thus, ³J_H4,H5 at 8.2 Hz indicates that H’s 4 and 5 are either close to antiperiplanar or synperiplanar, while the ³J_H3ax,H4, ³J_H3eq,H4, and³ J_H5,H6 coupling constants of 6.1, 3.2, and 2.1 Hz, respectively, exclude axial orientations for both H4 and H5. This ensemble of information points to a chair-like conformation with axial substituents at C4 and C5, with twisting of the C4-C5 bond to minimize repulsion between the hexafluoroisopropyl groups, which forces the near-eclipsing of H’s 4 and 5. In terms of spatial proximity, nOe correlations were observed between the 4-O-hexafluoroisopropyl proton and the ring protons H4 and H5. One of the two trifluoromethyl groups in the same 4-O-hexafluoroisopropyl ether also showed heteronuclear nOe correlations with H4 in the HOESY (¹H-¹⁹F) spectrum further supporting the assigned conformation (Figure 3).

Figure 3.

Nuclear Overhauser Correlations Establishing the Structure of 24.

Discussion

With reported pKa’s in aqueous solution varying between 4.53¹⁶ and 7.39^{17, 18} N-hydroxybenzotriazole performs similarly to acetic acid and delivers a moderate yield of the direct substitution product 13 with retention of configuration (Table 1, entry 1). The situation with N-hydroxyphthalimide (Table 1, entry 2) on the other hand is more complex: the direct substitution product 16, expected to be the major product on the basis of the pKa (6.1) of the nucleophile,¹⁹ was isolated in 36% yield. On the other hand, the double substitution product 17, isolated in 32% yield, approximates more closely to the reactivity pattern observed with phenol (4, Scheme 2) and was not anticipated. Clearly, other factors intervene with N-hydroxyphthalimide to promote elimination of the acetoxy group from the 4-position giving rise to the vinyl diazonium ion 10. Whatever the reason for the elimination, Michael addition of N-hydroxyphthalimide to 10 takes place under thermodynamic control as with phenol to give an equatorial adduct. Also as with phenol we propose that a second molecule of N-hydroxyphthalimide then protonates the diazoalkane axially on the α-face leading to a contact ion pair. However, unlike the phenol case where this contact ion pair collapses with loss of nitrogen and formation of the axial C-O bond, this ion pair containing a more stable anion follows the pattern established with acetic acid as nucleophile and lives sufficiently long for participation by the ring oxygen leading ultimately to nucleophilic substitution on the β-face with overall retention of configuration at the 5-position (Scheme 5).

Scheme 5.

Reaction of N-Hydroxyphthalimide with Vinyl Diazonium Ion 10

Acetohydroxamic acid, with a pKa of 9.40,¹¹ favors the diazoalkane 7 over its conjugate acid, the diazonium ion 6, leading to the formation of the vinyl diazonium ion 10. Conjugate addition then occurs on the β-face leading to a kinetic adduct, which undergoes intramolecular proton transfer leading to a zwitterion followed by collapse with loss of nitrogen to give the observed cis-fused product 18 (Table 1, entry 3, Scheme 6) in close overall analogy to the mechanism postulated for the use of β-naphthol as nucleophile and leading to the formation of 5.⁷ Turning to the use of N-(4-methoxybenzyl)acetohydroxamic acid, with an expected pKa similar to that of acetohydroxamic acid itself,¹¹ no substitution products were isolated, rather the alkene 19 was observed for the first time (Table 1, entry 4). We speculate that this formal reduction product arises from competing nucleophilic attack of the bulky nucleophile of the terminal nitrogen of the vinyl diazonium ion leading to a complex intermediate. This intermediate presumably decomposes with loss of nitrogen and of an unstable acyl nitronium ion, resulting in the formation of a vinyl anion ready for protonation to give alkene 19 (Scheme 6). Although N-Boc acetohydroxamic acid can be expected to have an acidity comparable to that of N-hydroxyphthalimide, it has considerable steric bulk and so might be expected to follow the same reaction path as N-(4-methoxybenzyl)acetohydroxamic acid resulting in the formation of alkene 19 rather than of any substitution product (Table 1, entry 5, Scheme 6).

Scheme 6.

Reaction of Hydroxamic Acids with Vinyl Diazonium Ion 10.

With p-nitrobenzaldehyde oxime (pKa 10.5)¹¹ as nucleophile the adduct 20, isolated in 26% yield (Table 1, entry 6), must arise from conjugate addition of the nucleophile to the vinyl diazonium ion 10, followed by protonation to the corresponding diazonium ion and explusion of nitrogen followed by deprotonation. Ketone 21 and, following elimination, α,β-unsaturated ketone 22 presumably arise by simple acid catalyzed hydrolysis of the relatively electron rich enol ether 20. With acetone oxime (pKa 12.4) the comparable adduct to 20 will be even more electron-rich and so does not withstand the reaction conditions; ultimately only a 38% overall yield of elimination products 14 and 15 could be isolated from this reaction mixture (Table 1, entry 7).

The aziridine 23 formed on the use of aniline as nucleophile is consistent with a mechanism involving kinetic attack of the nucleophile on the β-face of the vinyl diazonium ion 10 followed by tautomerization and cyclization to afford a triazoline (Scheme 7). Alternatively, by analogy with the well-known reaction of arene diazonium ions with amines,^{20, 21} the triazoline may be formed by initial attack of the amine onto the terminal nitrogen position of the diazonium moiety to give a vinyl triazene, followed by tautomerization and ring closure. The formation of an intermediate triazoline is supported by examination of the crude reaction mixture by ESI mass spectrometry, which revealed a major ion of m/z 530 (M+Na) that diminished over time in favor of the aziridine (m/z 501, M+Na). Loss of nitrogen and formation of the corresponding aziridines in this manner is the standard mode of decomposition of 1,2,3-triazolines. This mechanism is further supported by the early work of Saalfrank and Ackerman who studied the reaction of primary amines with simple β-ethoxyvinyl diazonium ions, leading to the isolation of 1,2,3-triazoles – with the intermediate triazoline undergoing elimination of the ethoxy group and aromatization, rather than loss of nitrogen and aziridine formation.^22–24

Scheme 7.

Mechanism of Triazoline Formation and Subsequent Decomposition to Aziridine 23

The attempted use of indole (pKa 16.2) as nucleophile gave a relatively clean reaction mixture from which was isolated a mixture of the two simple elimination products 14 and 15 in the unusually high yield of 80%. We suggest that this is the consequence of stabilization of the diazonium ion 6 in the form of an unstable triazene that retards elimination to the vinyl diazonium ion and ultimately decomposes with formation of the observed alkenes (Scheme 8). Related N-arylazoindoles have been isolated previously from the reaction of indole with arene diazonium salts under basic conditions.²⁵

Scheme 8.

Proposed stablization of Diazonium ion 6 by Triazene Formation

Finally, albeit the attempted use of 3,5-dimethylpyrazole as nucleophile in hexafluoroisopropanol afforded a complex reaction mixture from which no pyrazole-containing products could be isolated, the bis(hexafluoroisopropanol) adduct 24 was secured in 9% yield. This product is unusual and worthy of comment as, while there are numerous literature examples of the trapping of carbenium ions by hexafluoroisopropanol, often with support from X-ray crystal structures,^26–31 the formation of 24 requires hexafluoroisopropanol (pKa 9.3)³² to act first as nucleophile in a conjugate addition to the vinyl diazonium ion 10. Thus, 24 is envisaged as the product of kinetic addition of hexafluoroisopropanol to the β-face of 10, axial protonation of the resulting diazoalkane on the α-face by a second molecule of hexafluoroisopropanol and collapse of the so-formed tight ion pair to afford 24 (Scheme 9).

Scheme 9.

Reaction of Hexafluoroisopropanol with the Vinyl Diazonium Ion 10

The conformation of 24, a flattened chair conformation with both hexafluoroiospropyl groups pseudoaxial, is presumably preferred because of unfavorable steric interactions between the hexafluoroisopropyl groups in alternate conformations that would enable them to benefit from the gauche effect. A similar effect is seen in the X-ray crystal structure of the formal glycol derivative 25 that adopts an antiperiplanar relationship of the two bulky ethers, even though this forces the two C-C bonds into a gauche relationship (Figure 4).²⁹

Figure 4.

Solid State Conformation of Diether 25

Conclusion

A variety of hydroxylamine derivatives and weakly basic amines have been examined as possible nucleophiles in the Zbiral-type deamination of N-nitroso-N-acetyl neuraminic acid. Depending on the nature of the nucleophile products arise either from direct substitution of the nitrosoamide moiety with retention of configuration, or from double substitution of the nitroamide and the vicinal acetoxy group. The double substitution products are considered to be formed by a multi-step process involving elimination of the acetoxy group to form a vinyl diazonium ion followed by Michael addition and finally displacement of nitrogen from the resulting diazonium ion. The configuration at C5 is a function of the nature of the protonation step, inter or intramolecular, of the diazoalkane and of the tightness of the so-formed ion pair, which again is a function of the acidity of the nucleophile. Thus, intermolecular protonation takes place in the axial direction to afford a contact ion pair of the counterion with the equatorial diazo leaving group. When the ion pair is tight, ie, with counterions arising from less acidic nucleophiles (eg, phenol, hexafluoroisopropanol) the ion pair collapses with inversion of configuration and formation of an axial bond to the nucleophile. On the other hand, with more stable counterions (acetate, benzotriazole-1-oxide, phthalimide-1-oxide, etc) a looser ion pair is formed, the ring oxygen participates in explusion of nitrogen and the product is formed with overall retention of configuration.

Experimental Section

General.

All reactions were performed using oven-dried glassware under an atmosphere of argon. All reagents and solvents were purchased from commercial suppliers and were used without further purification unless otherwise specified. Chromatographic purifications were performed on silica gel (230–400 mesh) columns (20–50 g) of silica gel per gram of crude compound). Reactions were monitored by analytical thin-layer chromatography on pre-coated glass backed plates (w/UV 254) and visualized by UV irradiation (254 nm) or by staining with 25% H₂SO₄ in EtOH or ceric ammonium molybdate (CAM) solution. Specific rotations were measured on an automatic polarimeter with a path length of 100 mm in the solvent specified. Concentrations are given in g/100 mL. High resolution mass spectra (HRMS) were recorded with an electrospray ionization (ESI) source coupled to a time-of-flight (TOF) mass analyzer or with an electron impact (EI) source coupled to a TOF mass analyzer. ¹H, ¹³C, ¹⁹F, spectra were recorded on a 400, 500 or 600 MHz spectrometer. NMR solvents were used without purification. Chemical shifts are given in ppm (δ) and coupling constants (J) are given in Hz. Multiplicities are given as singlet (s), broad singlet (br s), doublet (d), triplet (t), doublet of doublets (dd), triplet of doublets (td), multiplet (m), apparent quartet (app q), apparent pentet (app p), etc.

General procedure for oxidative deamination.

Using the quantities described in the individual experiments, sodium 2,2,2-trifluoroethoxide and 18-crown-6 were dissolved in anhydrous CH₂Cl₂ under Ar and cooled to −10 °C. The solution was added to the nitrosyl sialoside (0.1 M solution in anhydrous CH₂Cl₂) at −10 °C under Ar. The mixture was stirred for 5 min at −10 °C. The nucleophile (5–20 equiv) dissolved in the solvent described under Ar at −10 °C was added to the reaction mixture in one portion. After stirring for 5 min, the reaction was quenched by addition of saturated NaHCO₃ solution and diluted with DCM. The reaction mixture was washed with NaOH/HCl (1M) to remove excess nucleophile. The organic layer was washed with brine, dried over anhydrous Na₂SO₄, and concentrated under reduced pressure to afford the crude product which was purified by column chromatography over silica gel.³³

Methyl (methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-3,5-dideoxy-D-glycero-β-D-galacto-non 2-ulopyranosid)onate (1).

Compound 1 (6 g, 95%) was obtained by a literature procedure⁴ over two steps as a white solid from N-acetylneuraminic acid (20 g, 64.7 mmol).

Methyl (methyl 4,7,8,9-tetra-O-acetyl-3,5-dideoxy-5-(anti/syn-N-nitrosoacetamido)-D-glycero-β-D-galacto-non-2-ulopyranosid)onate (2).

A solution of compound 1 (330 mg, 0.7 mmol) in dry dichloromethane (7 mL) was treated with dry pyridine (0.5 mL, 6.5 mmol, 10 equiv) and cooled to −10 °C. After stirring for 15 min, crushed nitrosyl tetrafluoroborate (382 mg, 3.0 mmol, 5 equiv) was added in one portion. The reaction mixture was stirred at −10 °C until TLC showed complete conversion (4–5 h). The mixture was diluted with cold dichloromethane (3 mL) and washed with cold 1N HCl, saturated NaHCO₃ and brine. The organic layer was dried over anhydrous Na₂SO₄ and concentrated under 10 °C to obtain 2 as a yellowish foam which was carried forward for next reaction without further purification⁴.

Methyl (methyl 4,7,8,9-tetra-O-acetyl-3-deoxy-5-O-(benzotriazol-1-yl)-D-glycero-β-D-galacto-non-2-ulopyranosid)onate (13).

The nitrosyl sialoside 2 (150 mg, 0.3 mmol) in CH₂Cl₂ (3 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (68 mg, 0.6 mmol), 18-crown-6 (74 mg, 0.3 mmol) in CH₂Cl₂ (2 mL) and HOBt (380 mg, 2.8 mmol, 10 equiv) in HFIP (2 mL) to afford 13 after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a colorless oil (78 mg, 48 %); ${\left[\alpha \right]}_{\text{D}}^{20}+45.7°$ (c 0.45, CH₂Cl₂).¹H NMR (600 MHz, CDCl₃) δ 7.97 (d, J = 8.4 Hz, 1H, ArH), 7.49 (d, J = 3.5 Hz, 2H, ArH), 7.36 (dt, J = 8.0, 3.8 Hz, 1H, ArH), 5.91 (dd, J = 5.0, 2.3 Hz, 1H, H7), 5.60 (ddd, J = 11.0, 9.4, 5.5 Hz, 1H, H4), 5.39 (ddd, J = 7.3, 5.0, 2.6 Hz, 1H, H8), 4.95 (t, J = 9.7 Hz, 1H, H5), 4.76 (dd, J = 12.5, 2.6 Hz, 1H, H9), 4.40 (dd, J = 10.1, 2.2 Hz, 1H, H6), 4.20 (dd, J = 12.4, 6.8 Hz, 1H, H9’), 3.80 (s, 3H, CH₃), 3.34 (s, 3H, CH₃), 2.60 (dd, J = 13.1, 5.5 Hz, 1H, H3e), 2.32 (s, 3H, CH₃), 2.11 (s, 3H, CH₃), 2.04 (s, 3H, CH₃), 1.67 (dd, J = 13.1, 11.2 Hz, 1H, H3a), 1.12 (s, 3H, CH₃). ¹³C{¹H} NMR (151 MHz, CDCl₃) δ 170.8, 170.6, 170.0, 169.3, 166.9, 143.3, 128.4, 127.9, 124.8, 120.3, 108.8, 98.7, 82.8, 71.2, 70.0, 69.3, 68.5, 62.3, 52.9, 51.8, 36.8, 21.1, 21.0, 20.9, 20.1. ESI-HRMS Calcd. for C₂₅H₃₁N₃NaO₁₃:([M+Na]⁺) m/z: 604.1755; found: 604.1756.

Methyl (methyl 4,7,8,9-tetra-O-acetyl-3-deoxy-5-O-(N-phthalimidoyl)-D-glycero-β-D-galacto-non-2-ulopyranosid)onate (16), Methyl (methyl 7,8,9-tri-O-acetyl-3-deoxy-4,5-di-O-(N-phthalimidoyl)-D-glycero-β-D-galacto-non-2-ulopyranosid)onate (17), Methyl (Methyl 4,7,8,9-Tetra-O-acetyl-3,5-dideoxy-β-D-arabino-non-4-en-2-ulopyranosid)onate (14), and Methyl (Methyl 4,7,8,9-Tetra-O-acetyl-3,5-dideoxy-β-D-ribo-non-5-en-2-ulopyranosid)onate (15).

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (90 mg, 0.7 mmol), 18-crown-6 (195 mg, 0.7 mmol) in CH₂Cl₂ (2 mL) and N-hydroxyphthalimide (1.2 g, 7.4 mmol, 20 equiv) to afford 16 after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a colorless oil (115 mg, 36%) and 17 as a colorless oil (102 mg, 27%, and an inseparable mixture of elimination products 14 and 15 as a colorless oil (1:2.5 ratio, 53 mg, 23%).

Compound 16: ${\left[\alpha \right]}_{\text{D}}^{20}-52.0°$ (c 0.4, CHCl₃). ¹H NMR (600 MHz, CDCl₃) δ 7.82 (dd, J = 5.5, 3.1 Hz, 2H, ArH), 7.75 (dd, J = 5.5, 3.1 Hz, 2H, ArH), 5.80 (dd, J = 5.3, 2.0 Hz, 1H, H7), 5.53 (ddd, J = 11.0, 9.0, 5.4 Hz, 1H, H4), 5.39 (ddd, J = 6.7, 5.3, 2.7 Hz, 1H, H8), 4.67 (dd, J = 12.4, 2.7 Hz, 1H, H9), 4.44 (t, J = 9.4 Hz, 1H, H5), 4.23 (p, J = 9.8, 2.1 Hz, 1H, H6), 4.16 (dd, J = 12.4, 6.7 Hz, 1H, H9’), 3.78 (s, 3H, CH₃), 3.29 (s, 3H, CH₃), 2.66 (dd, J = 13.2, 5.4 Hz, 1H, H3e), 2.25 (s, 3H), 2.10 (s, 3H, CH₃), 2.04 (s, 3H, CH₃), 1.72 (s, 3H, CH₃), 1.69 (dd, J = 13.1, 11.0 Hz, 1H, H3a). ¹³C{¹H} NMR (151 MHz, CDCl₃) δ 170.8, 170.5, 170.2, 169.4, 167.3, 163.3, 134.8, 128.9, 123.7, 98.6, 79.5, 71.0, 70.2, 69.4, 68.9, 62.4, 52.8, 51.7, 36.4, 21.1, 21.1, 20.9, 20.8. ESI-HRMS Calcd. for C₂₇H₃₁NNaO₁₅:([M+Na]⁺) m/z: 632.1591; found: 632.1592.

Compound 17: ${\left[\alpha \right]}_{\text{D}}^{20}-32.0°$ (c 0.25, CHCl₃). ¹H NMR (600 MHz, CDCl₃) δ 7.79 (dd, J = 5.5, 3.1 Hz, 2H, ArH), 7.73 – 7.68 (m, 4H, ArH), 7.67 (dd, J = 5.5, 3.0 Hz, 2H, ArH), 5.79 (dd, J = 5.7, 2.1 Hz, 1H, H7), 5.39 (td, J = 6.2, 2.7 Hz, 1H, H8), 5.17 (ddd, J = 10.5, 8.0, 5.5 Hz, 1H, H4), 4.86 (dd, J = 9.6, 7.9 Hz, 1H, H5), 4.67 (dd, J = 12.4, 2.7 Hz, 1H, H9), 4.23 (dd, J = 9.6, 2.1 Hz, 1H, H6), 4.16 (dd, J = 12.4, 6.5 Hz, 1H, H9’), 3.80 (s, 3H, CH₃), 3.23 (s, 3H, CH₃), 2.50 (dd, J = 13.4, 5.5 Hz, 1H, H3e), 2.32 (s, 3H, CH₃), 2.13 (dd, J = 13.4, 10.5 Hz, 1H, H3a), 2.10 (s, 3H, CH₃), 2.03 (s, 3H, CH₃). ¹³C{¹H} NMR (151 MHz, CDCl₃) δ 170.9, 170.4, 170.2, 167.4, 163.6, 163.3, 134.7, 134.3, 129.3, 128.8, 123.8, 123.6, 98.9, 82.5, 78.8, 70.8, 69.2, 68.6, 62.3, 53.0, 51.5, 35.5, 21.2, 20.9, 14.3. ESI-HRMS Calcd. for C₃₃H₃₂N₂NaO₁₆:([M+Na]⁺) m/z: 735.1650; found: 735.1650.

Compounds 14 and 15 (1:2.5 mixture) had NMR data consistant with the literature.⁷

Methyl (methyl 7,8,9-tri-O-acetyl-3-deoxy-4-O,5-O-(ethanimin-1-yl-N-yl)-D-glycero-β-D-talo-non-2-ulopyranosid)onate (18).

The nitrosyl sialoside 2 (200 mg, 0.4 mmol) in CH₂Cl₂ (4 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (90 mg, 0.7 mmol), 18-crown-6 (195 mg, 0.7 mmol) in CH₂Cl₂ (3 mL) and acetoxyhydroxamic acid (555 mg, 7.4 mmol, 20 equiv) to afford 18 after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a colorless oil (62 mg, 36%) and the single elimination product 15 as a colorless oil (46 mg, 28%). Compound 18; ${\left[\alpha \right]}_{\text{D}}^{20}-15.3°$ (c 0.85, CHCl₃) ¹H NMR (400 MHz, CDCl₃) δ 5.51 (dd, J = 5.6, 2.6 Hz, 1H, H7), 5.34 (td, J = 6.2, 2.6 Hz, 1H. H8), 4.57 (dd, J = 12.5, 2.7 Hz, 1H, H9), 4.33 (dd, J = 10.0, 2.6 Hz, 1H, H6), 4.22 (dd, J = 12.5, 6.3 Hz, 1H, H9’), 4.17 (dd, J = 10.0, 2.8 Hz, 1H, H5), 4.04 (dt, J = 4.2, 2.6 Hz, 1H, H4), 3.78 (s, 3H, CH₃), 3.23 (s, 3H, CH₃), 2.53 (dd, J = 15.5, 2.5 Hz, 1H, H3), 2.14 (s, 3H, CH₃), 2.10 (dd, J = 15.5, 4.2 Hz, 1H, H3’), 2.07 (s, 3H, CH₃), 2.03 (s, 3H. CH₃), 1.92 (s, 3H, CH₃). ¹³C{¹H} NMR (101 MHz, CDCl₃) δ 170.7, 170.3, 169.6, 167.9, 155.2, 97.7, 77.5, 77.2, 76.8, 70.5, 69.1, 68.8, 66.8, 66.1, 62.1, 52.8, 51.8, 34.9, 21.1, 20.8, 20.8, 17.4. ESI-HRMS Calcd. for C₁₉H₂₇NNaO₁₂:([M+Na]⁺) m/z: 484.1431; found: 484.1400.

Methyl (methyl 7,8,9-tri-O-acetyl-3,4,5-trideoxy-β-D-arabino-non-4-en-2-ulopyranosid)onate (19).

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and N-(4-methoxybenzyl)acetyl hydroxamic caid (975 mg, 5 mmol, 10 equiv) in CH₂Cl₂ (5 mL) to afford 19 after flash column chromatography over silica gel eluting with hexane/ethyl acetate (3:1), as a colorless oil (41 mg, 21%) and an inseparable mixture of elimination products 14 and 15 as a colorless oil (40 mg, 18%). ${\left[\alpha \right]}_{\text{D}}^{20}-10.1°$ (c 1.3, CHCl₃). ¹H NMR (400 MHz, CDCl₃) δ 5.82 (ddt, J = 10.0, 4.9, 2.3 Hz, 1H, H4), 5.63 (ddt, J = 10.5, 3.0, 1.5 Hz, 1H, H5), 5.45 (td, J = 6.2, 2.3 Hz, 1H, H8), 5.37 (dd, J = 6.2, 2.7 Hz, 1H, H7), 4.61 (dd, J = 12.5, 2.3 Hz, 1H. H9), 4.43 (tdq, J = 4.9, 2.6, 1.5, 0.9 Hz, 1H, H6), 4.22 (dd, J = 12.5, 6.1 Hz, 1H, H9’), 3.80 (s, 3H, CH₃), 3.28 (s, 3H, CH₃), 2.40 (ddt, J = 17.8, 4.0, 2.6 Hz, 1H, H3), 2.32 (dddd, J = 17.8, 4.8, 3.2, 1.4 Hz, 1H, H3’), 2.06 (s, 6H, 2CH₃), 2.04 (s, 3H, CH₃). ¹³C{¹H} NMR (101 MHz, CDCl₃) δ 170.8, 170.1, 170.0, 169.0, 123.8, 123.8, 97.6, 71.3, 70.3, 69.2, 62.5, 52.6, 51.5, 32.1, 21.0, 20.9, 20.8.ESI-HRMS Calcd. for C₂₇H₂₄NaO₁₀:([M+Na]⁺) m/z: 411.1262; found: 411.1248.

Deamination of nitrosyl sialoside 2 with N-(^tButyloxycarbonyl)acetyl hydroxamic acid as nucleophile. Methyl (7,8,9-tri-O-acetyl-3,5-dideoxy-β-D-arabino-non-4-oxo-2-ulopyranosid)onate (21) and Methyl (7,8,9-tri-O-acetyl-2,3,5-trideoxy-β-D-arabino-non-2-en-4-oxo-2-ulopyranosid) onate (22).

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and N-(^tbutyloxycarbonyl)acetyl hydroxamic acid (525 mg, 3 mmol, 6 equiv) in CH₂Cl₂ (5 mL) to afford 19 after flash column chromatography over silica gel eluting with hexane/ethyl acetate (3:1), as a colorless oil (20 mg, 10%), an inseparable mixture of elimination products 14 and 15 as a colorless oil (91 mg, 41%) and another inseparable mixture of compounds 21 and 22 (1.27:1 ratio, 35 mg, 17%) as a colorless oil.

Compound 22;^{7 1}H NMR (400 MHz, CDCl₃) δ 6.24 (d, J = 1.1 Hz, 1H, H3), 5.39 (ddd, J = 7.9, 5.3, 2.5 Hz, 1H, H8), 5.38 (dd, J = 7.9, 2.8 Hz, 1H, H7), 4.71 (ddd, J = 13.3, 4.1, 2.7 Hz, 1H, H6), 4.49 (dd, J = 12.5, 2.3 Hz, 1H, H9), 4.25 (m, 1H, H9), 3.87 (s, 3H, CH₃), 2.57 (dd, J = 16.9, 13.7 Hz, 1H, H5), 2.48 (dd, J = 16.9, 3.8 Hz, 1H, H5’), 2.14 (s, 3H, CH₃), 2.07 (s, 3H, CH₃), 2.06 (s, 3H, CH₃). ¹³C{¹H} NMR (101 MHz, CDCl₃) δ 191.8, 170.7, 170.0, 161.7, 158.3, 109.5, 77.7, 77.4, 77.2, 76.9, 70.1, 69.5, 61.7, 53.4, 37.8, 21.0, 20.8, 20.6.

Compound 21 was identified in the mixture by the following diagnostic signals; ¹H NMR (400 MHz, CDCl₃) δ 5.35 (ddd, J = 6.0, 5.7, 2.5 Hz, 1H, H8), 5.29 (dd, J = 6.0, 3.1 Hz, 1H, H7), 4.58 (dd, J=12.5, 2.6 Hz, 1H, H9), 4.25 (m, 1H, H6), 4.21 (dd, J = 12.5, 5.8 Hz, 1H, H9’), 3.82 (s, 3H, CH₃), 3.24 (s, 3H, CH₃), 2.74 (d, J = 14.9 Hz, 1H, H3), 2.61 (d, J = 14.9 Hz, 1H, H3’), 2.39 (br s, 1H, H5), 2.37 (br s, 1H, H5’), 2.13 (s, 3H, CH₃), 2.05 (s, 3H, CH₃), 2.04 (s, 3H, CH₃). ¹³C {¹H} NMR (101 MHz, CDCl₃) δ 201.6, 170.7, 170.2, 169.8, 167.2, 100.2, 71.3, 70.2, 69.0, 62.0, 53.0, 51.5, 48.0, 42.1, 21.0, 20.8, 20.8.

Methyl (methyl 7, 8, 9-tri-O-acetyl-3,5-dideoxy-4-O-(4-nitrophenylmethanimin-N-yl)-β-D-arabino-non-4-en-2-ulopyranosid)onate (20).

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and 4-nitrobenzaldehyde oxime (1.66 g, 10 mmol, 20 equiv) in THF (5 mL) to afford 20 after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a colorless oil (72 mg, 26%), an inseparable mixture of elimination products 14 and 15 as a colorless oil (53 mg, 24%) and another inseparable mixture of compounds 21 and 22 (17 mg, 8%) as a colorless oil. ${\left[\alpha \right]}_{\text{D}}^{20}-15.5°$ (c 0.9, CHCl₃). ¹H NMR (600 MHz, CDCl₃) δ 8.22 (d, J = 8.8 Hz, 2H, ArH), 8.02 (s, 1H, NCH), 7.73 (d, J = 8.8 Hz, 2H, ArH), 5.57 (ddd, J = 7.3, 4.5, 2.8 Hz, 1H, H8), 5.53 (dt, J = 4.6, 1.2 Hz, 1H, H7), 5.20 (d, J = 4.2 Hz, 1H, H5), 4.87 (dt, J = 6.4, 5.0 Hz, 1H, H4), 4.42 (dd, J = 12.2, 2.8 Hz, 1H, H9), 4.36 (dd, J = 12.1, 7.4 Hz, 1H, H9’), 3.74 (s, 3H, CH₃), 3.39 (s, 3H, CH₃), 2.56 (dd, J = 14.0, 4.8 Hz, 1H, H3), 2.25 (dd, J = 14.0, 5.2 Hz, 1H, H3’), 2.15 (s, 3H, CH₃), 2.07 (s, 3H, CH₃), 2.06 (s, 3H, CH₃). ¹³C{¹H} NMR (151 MHz, CDCl₃) δ 170.9, 170.1, 169.4, 167.8, 149.7, 148.6, 147.2, 138.2, 127.8 (2 Carbons), 124.1(2 Carbons), 99.4, 98.5, 72.3, 70.5, 70.5, 62.0, 52.7, 52.3, 34.7, 21.0, 20.9 (2 Carbons). ESI-HRMS Calcd. for C₂₄H₂₈N₂NaO₁₃:([M+Na]⁺) m/z: 575.1484; found: 575.1486.

Deamination of nitrosyl sialoside 2 with acetone oxime as nucleophile.

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and acetone oxime (731 mg, 10 mmol, 20 equiv) in CH₂Cl₂ (5 mL) to afford 14 and 15, after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a colorless oil (84 mg, 38%).

Methyl (methyl 7,8,9-tri-O-acetyl-3,4,5-trideoxy-4-N,5-N-phenyliminyl-D-glycero-β-D-talo-non-2-ulopyranosid)onate (23).

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and aniline (0.83 mL, 9.2 mmol, 20 equiv) to afford 23, after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a yellowish oil (110 mg, 49%). ${\left[\alpha \right]}_{\text{D}}^{20}-45.0°$ (c 0.5, CH₂Cl₂).¹H NMR (600 MHz, CDCl₃) δ 7.22 (dd, J = 8.5, 7.3 Hz, 2H, ArH), 6.98 – 6.94 (m, 3H, ArH), 5.69 (dd, J = 6.0, 3.9 Hz, 1H, H7), 5.52 (td, J = 5.9, 2.8 Hz, 1H, H8), 4.59 (dd, J = 12.5, 2.8 Hz, 1H, H9), 4.32 (dd, J = 12.5, 5.9 Hz, 1H, H9’), 4.16 (t, J = 3.6 Hz, 1H, H6), 3.80 (s, 3H, CH₃), 3.22 (s, 3H, CH₃), 2.51 (dd, J = 7.1, 3.4 Hz, 1H, H5), 2.47 (ddd, J = 7.1, 5.3, 1.7 Hz, 1H, H4), 2.37 (dd, J = 15.3, 1.8 Hz, 1H, H3pe), 2.26 (dd, J = 15.3, 5.3 Hz, 1H, H3pa), 2.15 (s, 3H, CH₃), 2.09 (s, 3H, CH₃), 2.08 (s, 3H, CH₃). ¹³C{¹H} NMR (151 MHz, CDCl₃) δ 170.8, 170.4, 170.2, 169.4, 153.4, 129.2, 123.1, 120.6, 97.6, 71.4, 70.3, 70.2, 62.2, 52.7, 52.0, 37.1, 35.3, 31.2, 21.1, 21.0, 20.9. ESI-HRMS Calcd. for C₂₃H₂₉NNaO₁₀:([M+Na]⁺) m/z: 502.1689; found: 502.1682.

Deamination of nitrosyl sialoside 2 with indole as nucleophile.

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and indole (1.17 g, 10 mmol, 20 equiv) in CH₂Cl₂ (5 mL) to afford 14 and 15, after flash column chromatography over silica gel eluting with hexane/ethyl acetate (1:1), as a colorless oil (182 mg, 81%).

Methyl (methyl 7,8,9-tri-O-acetyl-3-deoxy-4,5-di-O-hexafluoroisopropyl-D-glycero-β-D-ido-non-2-ulopyranosid)onate (24).

The nitrosyl sialoside 2 (267 mg, 0.5 mmol) in CH₂Cl₂ (5 mL) was deaminated using the general procedure for oxidative deamination with sodium 2,2,2-trifluoroethoxide (122 mg, 1 mmol), 18-crown-6 (291 mg, 1.1 mmol) in CH₂Cl₂ (2.5 mL) and 3,5-dimethylpyrazole (960 mg, 10 mmol, 20 equiv) dissolved in HFIP (5 mL) to afford 24, after flash column chromatography over silica gel eluting with hexane/ethyl acetate (3:1), as a colorless oil (31 mg, 9%). ${\left[\alpha \right]}_{\text{D}}^{20}-17.5°$ (c 1.5, CH₂Cl₂). ¹H NMR (600 MHz, CDCl₃) δ 5.24 (td, J = 6.0, 3.4 Hz, 1H, H8), 5.10 (p, J = 6.0 Hz, 1H, CH), 5.03 (t, J = 5.5 Hz, 1H, H7), 4.75 (p, J = 6.0 Hz, 1H, CH), 4.34 (dd, J = 12.3, 3.4 Hz, 1H, H9), 4.17 (dd, J = 12.3, 6.2 Hz, 1H, H9’), 3.80 (s, 3H, CH₃), 3.71 (ddd, J = 8.7, 6.0, 3.2 Hz, 1H, H4), 3.39 (s, 3H, CH₃), 3.11 (dd, J = 8.2, 2.1 Hz, 1H, H5), 3.02 (dd, J = 5.3, 2.1 Hz, 1H, H6), 2.49 (dd, J = 15.9, 6.1 Hz, 1H, H3a), 2.38 (dd, J = 15.9, 3.2 Hz, 1H, H3e), 2.11 (s, 3H, CH₃), 2.08 (s, 3H, CH₃), 2.05 (s, 3H, CH₃). ¹³C{¹H} NMR (151 MHz, CDCl₃) δ 170.6, 170.0, 169.7, 166.7, 101.1, 78.4, 73.5, 73.2, 73.0, 72.8, 72.6, 70.5, 69.8, 69.6, 69.4, 69.2, 68.9, 68.7, 61.6, 58.4, 53.2, 53.0, 52.7, 36.1, 20.7, 20.7. ¹³C{¹⁹F,¹H} NMR (126 MHz, CDCl₃) δ 170.7, 170.0, 169.7, 166.7, 121.7, 121.5, 121.3, 121.2, 101.2, 78.5, 73.1, 70.5, 69.8, 69.2, 61.6, 58.4, 53.3, 53.1, 52.7, 36.2, 20.8, 20.7. ¹⁹F NMR (471 MHz, CDCl₃) δ −75.42 (q, J = 8.0, 7.1 Hz), −75.47 – −75.59 (m), −76.46 (h, J = 8.5, 7.3 Hz), −76.80 (qd, J = 9.2, 6.0 Hz). ESI-HRMS Calcd. for C₂₃H₂₆F₁₂NaO₁₂:([M+Na]⁺) m/z: 745.1130; found: 745.1125.

Preparation of 26 by reduction of 13.

Compound 13 (22 mg, 0.04 mmol) was dissolved in anhydrous THF (0.2 mL), and treated at room temperature with glacial acetic acid (0.8 mL) and then Zn-dust (50 mg). The reaction mixture was heated to reflux with stirring until completion (2 h) before cooling to room temperature, quenching with water and filtration through Celite. The filtrate was washed with water and brine, dried over anhydrous Na₂SO₄ and concentrated under reduced pressure. The crude mixture was then acetylated using acetic anhydride (1 mL) and pyridine (0.5 mL) at room temperature, concentrated, and purified by column chromatography over silica gel eluting with (hexane/ ethyl acetate 1:1) to afford 26 as a white foam (22 mg, 97%).