Synthesis of Novel O-Acylated-D-ribono-1 , 5-lactones and Structural Assignment Supported by Conventional NOESY-NMR and X-ray Analysis

Este trabalho apresenta um método simples para a caracterização estrutural de 3,4-O-benzilideno-D-ribono-1,5-lactonas e análogos, fundamentado em técnicas convencionais de RMN e experimentos de NOESY em solução. 2-O-Acil-3,4-O-benzilideno-D-ribono-1,5-lactonas foram preparadas em bons rendimentos a partir da acilação das lactonas de Zinner empregando cloretos de ácido sob condições básicas. A estrutura de 2-O-(4-nitrobenzoil)-3,4-O-benzilidenoD-ribono-1,5-lactona foi determinada por difração de raios-X e confirmou, de forma inequívoca, os dados espectroscópicos obtidos.


Introduction
][10] The chemical complexity of carbohydrate derivatives, due to subtle differences in reactivity of the various hydroxyl groups and the possible formation of species co-existing in equilibrium under different reaction conditions, may lead to unexpected processes ranging from rearrangements to functional group migrations. 11,12In many cases, single crystal X-ray analysis is the only reliable method for correct structural and conformational assignments. 13,14Aldonolactones 15 (modified sugars with the anomeric centre in its higher oxidation state) have been widely employed in synthesis as an alternative to simple carbohydrates.D-Ribono-1,4-lactone (D-ribono--lactone, 1) is one of the most useful chiral precursors from this class, [16][17][18][19] although the issues associated with product characterization due to unpredictable transformations are also present to a great extent.For instance, the product obtained from the reaction of D-ribono-1,4-lactone (1)  with benzaldehyde in acidic medium, which was initially assigned as being 3,5-O-benzylidene-D-ribono-1,4-lactone (2), 20,21 had the correct structure unequivocally established as 3,4-O-benzylidene-D-ribono-1,5-lactone (3) by single crystal X-ray diffraction analysis of its O-acetyl derivative (Scheme 1). 22A complex rearrangement involving acetal migration and ring expansion sequences has been proposed for this transformation. 10,14][30] In our ongoing studies on synthetic transformations of nucleosides and carbohydrates as inhibitors of trypanosomal gGAPDH enzyme, [31][32][33] the synthesis of selectively acylated acetals of D-ribono-1,5-lactone 3 was required for screening programs.Based on the above statements and bearing in mind that derivatives of O-benzylidene-D-ribono-1,5-lactone 3 undergo acid-catalyzed deprotection with subsequent rearrangement to the corresponding D-ribono- 1,4-lactones, 13 we anticipated that any further chemical transformation on 3 should not be merely assumed without proper spectroscopic support. 34,35In this paper we describe a simple and efficient method for structural assignment of benzylidene-D-ribono-1,5-lactones by 1 H NMR experiments involving NOESY measurements in solution.

Results and Discussion
2-O-Acyl-D-ribono-1,5-lactones 4 were prepared in high yield and gram amounts by acylation of 3 with acyl chlorides in excess pyridine followed by aqueous workup and crystallization (Scheme 2).The experimental procedure is straightforward and all reagents are easily available.It should be noted that application of alternative methods for acylation employing harmless reagents was severely restricted due to the poor solubility of 3 in many ordinary solvents.
Unequivocal structural determination for lactone 4c was done by single crystal X-ray methods after a careful recrystallization of the solid product in acetone/ethanol.The pyranose ring adopts a boat-like conformation in the solid state, with the nitrobenzoyl group being positioned in a pseudo-equatorial orientation in order to minimize steric repulsions from the adjacent 3,4-dioxolanyl moiety (Figure 1).These results confirm that rearrangements coming from acyl migration, acetal isomerization or ring expansioncontraction did not operate under the stated reaction conditions for acylation of D-ribono-1,5-lactone 3. 14,22,24 Scheme 2. Scheme 3.Although the acylated D-ribono-1,5-lactone 4c was the only crystalline compound suitable for X-ray determination, a complete characterization of D-ribono-1,5-lactone 4 was accomplished by NMR spectroscopic data.Analysis by 13 C NMR spectroscopy has been previously proposed as a useful tool to establish the size of the lactone ring, supported by considerable differences in the chemical shifts found for C-3 and C-4 in a few lactones studied. 13However, a broader data compilation for 1,4-and 1,5-lactones revealed that no practical correlation could be rationalized (Table 2).
After searching for general trends in the 1 H NMR to distinguish among several related sugar lactones, studies dealing with coupling constants ( 3 J) revealed some useful patterns (Table 3).Small coupling between H-2 and H-3 were found for D-ribono-1,5-lactones 3-7 ( 3 J 2,3 1.5-3.4Hz) in comparison with those for D-ribono-1,4-lactones 1,8-10 ( 3 J 2,3 = 5.3-6.2Hz).On the other hand, high values were encountered for H-3, H-4 coupling in D-ribono-1,5-lactones 3-7 ( 3 J 3,4 7.9-8.2Hz), while they are nearly absent in D-ribono-1,4-lactones 1,8-10 ( 3 J 3,4 <1 Hz).It is also worth noting that the very low values assigned for coupling between H-4 and H-5a/b in D-ribono-1,5-lactones 3-7 ( 3 J 4,5 <1.7 Hz), compared with those for D-ribono-1,4lactones 1,8-10 ( 3 J 4,5 2.4-4.5 Hz), are probably associated with an adopted gauche conformation in solution with dihedral angles close to 60 o , leading to weak spin-spin interactions as expected by the Karplus-Conroy equation. 9,38e NOESY experiments performed with selected lactones not only confirmed the results deduced from coupling constant analysis but also revealed other important structural features (Table 4).Besides the anticipated presence of cross-peaks between H-3 and H-4 in the D-ribono-1,5-lactones 4a-c and 7 (H-3 and H-4 are syn), and its absence in D-ribono-1,4-lactones 9 and 10 (H-3 and H-4 are anti), long-range contacts between H-2 and H-5b were also observed for all ribono-1,5-lactones but not for 1,4-lactones.Although no reasonable correlation was noted between H-4 and H-5a/b in D-ribono-1,5-lactones 4a-c and 7, this observation was expected based on the  H NMR spectra (400 MHz) were acquired using DMSO-d 6 as the solvent (unless otherwise specified), but no significant effect over the chemical shifts or coupling constants was observed when other common deuterated solvents such as CDCl 3 and C 6 D 6 were employed; b Coupling constants were not detected in the NMR spectrum; c These assignments may have to be reversed; d Data obtained in CDCl 3 as the solvent; e Data obtained in C 6 D 6 as the solvent; f Values could not be assigned due to complex multiplicities found for H-4 and H-5.C NMR spectra (100 MHz) were acquired using DMSO-d 6 as the solvent (unless otherwise specified), but no significant effect over the chemical shifts or coupling constants was observed when other common deuterated solvents such as CDCl 3 and C 6 D 6 were employed; b Data obtained in CDCl 3 as the solvent; c These assignments may have to be reversed.

Conclusions
Structural assignments for O-benzylidene-D-ribono-1,5-lactones were achieved using conventional NMR/ NOESY techniques and were confirmed by X-ray crystallography.In a general sense, the observed correlation between H-3 and H-4 is characteristic for D-ribono-1,5- lactones, but not for D-ribono-1,4-lactones.In addition, significant spin-spin interaction between H-4 and H-5 are present in D-ribono-1,4-lactones, while they are absent in D-ribono-1,5-lactones.The particular trends featured by both D-ribono-1,4-and D-ribono-1,5-lactones can be helpful resources to account for their physicochemical characterization in a simple and reliable way.

General procedure for the preparation of 2-O-acyl-Dribono-1,5-lactones (4a-d)
To a solution containing 0.20 g (0.85 mmol) of Zinner's lactone (3) in 1.0 mL of anhydrous chloroform and 1.0 mL of dry pyridine under argon at 25 o C was added 1.3 mmol (1.5 equiv.) of the corresponding acyl chloride.After stirring for 15-18 h, the reaction mixture was diluted with CH 2 Cl 2 and successively washed with 5% HCl, 5% NaHCO 3 , and H 2 O.The organic extract was dried over Na 2 SO 4 , filtered, and concentrated under reduced pressure to give solid products that were purified by crystallization in acetone/ethanol (4a-c) or ethyl acetate (4d).

Single-crystal X-ray structure determinations
Low temperature X-ray diffraction data collection ( scans and scans with offsets) of 4c was performed on an Enraf-Nonius Kappa-CCD diffractometer (95 mm CCD camera on -goniostat) using graphite-monochromated MoK radiation (0.71073 Å).Data were collected up to 50° in 2 , with a redundancy of 4. The final unit cell parameters were based on all reflections.Data collections were made using the COLLECT program, 39 integration and scaling of the reflections were performed with the HKL Denzo-Scalepack system of programs. 40No absorption correction was applied.The structure was solved by direct methods with SHELXS-97. 41The model was refined by full-matrix least squares on F 2 with SHELXL-97. 42All the hydrogen atoms were stereochemically positioned and refined with the riding model. 42Hydrogen atoms were set isotropic with a thermal parameter 20% greater than the equivalent isotropic displacement parameter of the atom to which each one was bonded.The programs SHELXS-97, 41 SHELXL-97 42 and ORTEP-3 43 were used within WinGX 44 to prepare materials for publication.