Synthesis and Antiacetylcholinesterase Activity of New D-Glyceraldehyde Heterocyclic Derivatives

Neste trabalho são apresentados os procedimentos para a síntese de diferentes compostos heterocíclicos a partir do 2,3-O-isopropilideno-D-gliceraldeído: ciclização intramolecular, cicloadição 1,3-dipolar e reação de acoplamento bimolecular. Os produtos sintetizados foram caracterizados por espectroscopia de H RMN e C RMN e análise elementar. Os novos heterocíclicos e seus derivados (12 compostos) foram testados como inhibidores da enzima acetilcolinesterase.


Introduction
Alzheimer´s disease (AD) is a progressive, degenerative, and irreversible disorder that causes intellectual impairment and cognitive dysfunction. 1 In the last decade, treatment for AD has been based on the "cholinergic hypothesis". 2This hypothesis suggested that patients with AD suffer from a deficit of cholinergic function in the brain such as decrease in hipocampal and cortical levels of acetylcholine (ACh) and associated enzyme choline transferase.Inhibition of acetylcholinesterase (AChE), an enzyme responsible for the metabolic break-down of ACh, can restore the level of ACh in the brain. 3Hence the development of drugs for more serious AD is essential.
In the search for new molecules with the potential effect of inhibiting the enzyme acetylcholinesterase it is important to note that many heterocycles have shown significant inhibitory activity on this enzyme, including three of the four AD drugs currently on the market. 4he series of heterocycles known as benzoisoxazoles have been developed as potent and selective inhibitors of the enzime AChE with IC 50 values between 0.8-14 nM. 5 In addition, ester derivatives of tetrahydroazocines were synthesized and evaluated as inhibitors of the enzyme AChE, showing to be selective to the enzyme with IC 50 values in the range of 5-40 mM. 6A series of tacrine analogues has also been synthesized and evaluated as inhibitors of acetylcholinesterase/butyrylcholinesterase and modulators of Ca +2 channels and nicotinic receptors. 7n the present study, a series of new D-glyceraldehyde heterocyclic derivatives were synthesized and were evaluated as potential inhibitors of acetylcholinesterase, so that current approaches to the treatment of cognitive and behavioral symptoms of AD make use of structurally diverse cholinesterase inhibitors.

Results and Discussion
The heterocyclic compounds and their derivatives were synthesized from 2,3-O-isopropylidene-D-glyceraldehyde using simple and efficient methodologies (Scheme 1).
Compound 3 was obtained using intramolecular heterocyclization as we have previously described. 8n turn compounds 5, 6 and 8 were synthesized by 1,3-dipolar cycloaddition from the 2,3-O-isopropylidene-D-glyceraldehyde oxime using alkynes 9 or alkenes 10 as dipolarophiles.The 1,2,4-oxadiazole derivatives (11 and  12) were obtained by cyclization of O-acyl amidoxime, which were synthesized using the technique described in the literature. 11Compounds 13-17 were obtained by deprotection of the corresponding precursors with acid medium.After purification by chromatographic column or recrystallization, compounds 2, 5-7, 11-14, 16 and 17 were obtained with moderate yields (see experimental) and were characterized by NMR spectroscopy.Spectroscopic data were in full agreement with those expected.
The syntheses of 3, 8 and 15 involved the formation of new asymmetric centre.Spectroscopic analysis of these compounds showed both diasteromers, in quasiequimolecular ratio.These facts can be explained because during the process of cyclization, intramolecular heterocyclization (compound 3) or 1,3-dipolar cycloaddition (compounds 8 and 15), there is no any chiral induction.
The compounds 3, 5-8 and 11-17 (Scheme1) were evaluated as potential inhibitors of the enzyme AChE.AChE inhibitory activity was measured at room temperature by the colorimetric method previously described. 12The results of enzymatic inhibition are shown in Table 1 expressly as IC 50 in mg mL -1 and mM.
All compounds showed moderate inhibitory activity of AChE, being compounds 3, 5 and 7 with IC 50 values of 110, 220 and 190 mM respectively.The most active compounds possess one of the following structural characteristics: a dioxolanic ring (3, 5 and 7), an acetate group (3), a larger number of carbon atoms (5, 7).

Conclusion
In summary, a new group of heterocyclic derivatives of 2,3-O-isopropylidene-D-glyceraldehyde with protected and unprotected hydroxyl groups were efficiently synthesized.The new derivatives were characterized by 1 H NMR, 13 C NMR and 2D heteronuclear techniques.Twelve compounds were evaluated as inhibitors of acetylcholinesterase showing moderate to low activity.The most active compounds were 3, 5 and 7 with IC 50 less than 220 mM .

Experimental
Optical rotations were recorded at 20 °C on a Perkin Elmer 343 polarimeter, and the melting points were uncorrected. 1H NMR and 13 C NMR spectra were recorded on a Bruker AC-200 spectrometer at 200 MHz and 50 MHz.Elemental analyses were performed by UMYMFOR, CONICET-University of Buenos Aires, Argentina.Analytical TLC was conducted on Silica Gel 60G (Merck) on precoated plates and visualization was made by UV light and ethanol/sulfuric acid (10:1) or cerium molibdate, followed by heating.Column-chromatographic separations were performed on Silica Gel (240-400 mesh, Merck).Solvents were reagent grade and, in most cases, dried and distilled before use according to standard procedures.

General procedure 2: deprotection of nucleoside analogues
A mixture of 2.04 mmol of 2,3-O-isopropylidene derivatives (5, 7, 8, 11, 12), 2 mL of TFA and 2 mL of water was stirred at room temperature for 4 h.The reaction mixture was evaporated and the deprotected compounds were purified as described in each cases.