Cyclization of 1,2:3,4-di-O -isopropylidene-  - D - galacto -1,6-hexodialdo-1,5-pyranose acylhydrazone and semicarbazone

Cyclization of 1,2:3,4-di-O -isopropylidene-  - D - galacto -1,6-hexodialdo-1,5-pyranose benzoylhydrazone using acetylating mixtures led us to the corresponding (2 R )- and (2 S )-5- phenyl-1,3,4-oxadiazoline derivatives. The same conditions applied to the semicarbazone produced the 5-methyl-1,3,4-oxadiazoline derivative as the main compound, which is formed with acetylating mixtures even at room temperature. X-Ray analysis and NMR techniques were used to determine the stereochemistry of the new asymmetric centers .


Introduction
The synthesis of 3-N-acyl-1,3,4-oxadiazolines is a well-known reaction and there is a lot of information on this field, however in recent years the number of publications has increased due to the great potential of this heterocyclic ring as chemotherapeutic agent. 1,2Nevertheless, some aspects of heterocyclization reaction are not well known.
Looking for new potential biological agents, we studied the differences between heterocyclization of acylhydrazone and semicarbazone obtained from protected carbohydrate derivatives.Also, a hypothesis for their different behavior is proposed.
Semicarbazone derivative 2 was synthesized by reaction of 1,2:3,4-di-O-isopropylidene--Dgalacto-1,6-hexodialdo-1,5-pyranose with semicarbazide hidrochloride and sodium bicarbonate in ethanolic solution.After removal of salts the crude product was obtained as a syrup, which crystallized from methanol.The 1 H NMR analysis performed on the syrup indicated that there is only one isomer, unlike benzoyl hydrazone 1 3 , which was synthesized as a mixture of anti:syn isomers in a 3:1 relationship.We performed a single crystal x-ray diffraction analysis of 2 and found an anti configuration for the C=N bond, but also presented a deviation respect to the expected zig-zag conformation.This orientation of the carbonyl group relative to the N=C group enables the formation of intermolecular hydrogen bonds, contributing to the ordering of the molecules and subsequent crystallization (Figure 1).Both molecules (2 and 3b) share their leftmost part (Figure 2, primed labels), and thus have a number of characteristics in common: in both units the fused dioxolane cycles prevent the pyranose ring from assuming a regular conformation; it exhibits instead a highly distorted twist-boat geometry clearly evidenced through the comparison of some selected torsion angles, viz., C2'-C3'-C4'-C5': 12.6(2)º (3b) or -3.8 (2) (implying that the C2'-C3' and C4'-C5' bonds are nearly eclipsed) vs. C2'-C1'-O1'-C5': 41.3(2)º (in 3b), 47.4(1)º (in 2) (showing that the C2'-C1' and O1'-C5' bonds are far from coplanar).
Compounds 3a and 3b were characterized spectroscopically, and a NOESY experiment was performed for 3a.In this experiment, the most relevant cross peak correlations were H4-H2(Het) and -CH3(C)-Ph, which confirmed the 2R stereochemistry for compound 3a.The observed correlations are depicted in Figure 3.When compound 2 was subjected to the same reaction conditions methyloxadiazolines 4a (R) and 4b (S) were obtained, although some byproducts were also detected.In order to perform a cleaner reaction, we carried out a microwave assisted synthesis, using the same reagents and heating at 140 °C during 60 minutes and we obtained 4a and 4b with better yields.Stereochemistry of compound 4a and 4b were determined by comparison with NMR data of compounds 3a and 3b.Conditions, yields, and R:S relationship for compounds 3 and 4 are shown on Table 1.After 15 min of irradiation, we observed the formation of both oxadiazolines (4a and 4b) and the presence of a third compound (5), which disappears after 60 min of irradiation.The chromatographic behavior of 5 is similar to that observed for the main byproduct of the thermal treatment.Once isolated, compound 5 was identified as 4-N-acetyl-1,2:3,4-di-O-isopropylidene--D-galacto-1,6-hexodialdo-1,5-pyranose semicarbazone.Hang et al. 4 proposed a mechanism for acylhydrazone cyclization via two different pathways, the direct and the indirect one.This latter route would generate the methyloxadiazoline as results of prolonged heating.When we treated semicarbazone 2 with acetic anhydride and pyridine at reflux, the product of direct cyclization was not observed and the main products were methyloxadiazolines (4a and 4b).When the same reaction was carried out at room temperature, the formation of compound 5 and 4b was observed within two hours of reaction.After 60 hs the reaction was stopped and we could isolate the oxadiazoline 4b (6%) along with traces of 4a, compound 5 (13%), and the unreacted compound 2 (35%); meanwhile N2-acetyl or N2,N4diacetyl derivatives were not detected.
These results demonstrated that the formation of methyloxadiazoline could not be attributed exclusively to the reaction temperature.Under these conditions, N4 would be the first acetylated nitrogen, since it is not sterically hindered, giving compound 5.The proposed reaction mechanism 4 (Scheme 2) starts with a N2 acylation, followed by a cyclization assisted by acetic anhydride.Based on this hypothesis, we propose that, if some of N2 acetylation takes place, this derivative (6) can rearrange assisted by the N4 electron pair, and undergo cyclization at low temperature.If we assume that the mechanism is similar to the one proposed for thiosemicarbazones cyclization, via carbocation intermediate 5 the result is the same.

Scheme 2. Proposed cylization mechanism for semicarbazones
According to our previous experiences on cyclization of sterically hindered thiosemicarbazones and acylhydrazones, 6 the stereochemistry of the molecule can rule the side of attack of sulfur atom, but this influence was not observed for oxygenated derivatives.Other authors reported that predominance of R or S oxadiazoline isomer is a direct consequence of the acylhydrazone syn:anti relationship, 4,7 but no R:S selectivity was observed for the oxadiazoline derivative because syn and anti isomers can be equilibrated by temperature or the presence of acetic acid.
During cyclization of 2 at room temperature, acetic anhydride with excess of pyridine was used, so, isomerization of anti form of 2 could not take place, and the main heterocyclic compound obtained was 4b.Taking into account that the starting material was a single isomer we can conclude that the preferential formation of 4b is a consequence of the stereochemistry of C=N bond on compound 2. On the other hand, some calculations using molecular mechanics, 8 showed that compound 4a is about 6 Kcal/mol more stable than 4b, so 4b must be a kinetic product meanwhile 4a must be the thermodynamic one.

Conclusions
Taking into account our experimental results, we can conclude that the cyclization of benzoylhydrazone 1 using acetic anhydride and pyridine at reflux, yielded 1,3,4-oxadiazoline derivatives (3a and 3b) as a mixture of syn:anti isomers.Under the same conditions, the semicarbazone 2 yields the methyloxadiazoline instead of the product of direct cyclization.This fact can be explained by the presence of the N4 electron pair, which promotes the loss of the carbamoyl group.Also, we performed the acetylation of compound 2 at room temperature and found that the cyclization takes place yielding preferentially 4b, which would be the kinetic reaction product.

Experimental Section
General.Elemental analysis was performed on an Exeter Analytical CE-440 elemental analyzer.Optical rotations were recorded at 20 °C on a Perkin Elmer 343 polarimeter. 1 H, 13 C NMR spectra were recorded on solution of CDCl3 on a Bruker AC-200 spectrometer, operating at 200, 50 MHz respectively; or a Bruker AMX-500 spectrometer, operating at 500, 125 MHz respectively.Assignments of the 1 H and 13 C NMR spectra were confirmed with the aid of two dimensional techniques 1 H, 13 C (COSY, HSQC).Microwave-assisted synthesis was performed on an Anton Paar Monowave 300 microwave reactor with external surface sensor in a sealed reaction vessel.Chromatographic purifications were performed by flash column with Merck silica gel 60.The chemicals used in this work purchased from Aldrich and were used without further purification.X-ray data was collected on an Oxford Diffraction Gemini CCD S Ultra single crystal diffractometer using Mo Ka radiation (λ = 0.71073 Å). 9 A multi-scan absorption correction was applied. 10The structure was solved by direct methods and refined by full-matrix least squares against F2 using all data. 11All non-H atoms were refined anisotropically, while H atoms were located at idealized positions with their displacement parameters riding on the values of their parent atoms.The general-purpose crystallographic program PLATON 12 was used for the structure analysis and presentation of the results.The figures were drawn with XP. 11
General synthetic procedure for thermal cyclization of compound 1 and 2. Compound 1 (0.352 g, 0.9 mmol) or 2 (0.399 g, 1.3 mmol) were dissolved in pyridine (2.5 mL) and acetic anhydride (2.5 mL).The resulting mixture was refluxed (see Table 1) and then was left to reach room temperature.Once cold, some ethanol was added and the reaction medium was evaporated at reduced pressure, giving a glassy residue.The residue was purified using flash dry column on Silicagel G with mixtures of cyclohexane/acetone and obtained compounds 3a and 3b or 4a and 4b.Microwave-assisted synthesis of compounds 4a and 4b.Compound 2 (0.4245 g, 1.3 mmol) were dissolved in pyridine (3.0 mL) in a microwave vessel and acetic anhydride (2.5 mL) was added.The sealed vessel with the mixture was heated at 140 °C with stirring (900 rpm) and the temperature was held for 60 min.Once cold, the mixture was transferred to a round bottom flask, some ethanol was added and the reaction medium was evaporated at reduced pressure, giving a glassy residue.The residue was purified as described previously and compounds 4a and 4b were obtained in a 68% yield (0.321 mg).

Figure 1 .
Figure 1.Drawing of the crystal packing of compound 2.

Figure 3 .
Figure 3. Cross correlations found in the NOESY experiment for compound 3a.

Table 1 .
Conditions for synthesis of compounds 3 and 4, yield, and R:S relationship.