Enantioselective Synthesis of both (-)-( R )-and ( + )-( S )-Angustureine Controlled by Enzymatic Resolution

Uma nova síntese dos enantiômeros (-)-(R)e (+)-(S)-angustureina, assim como do racemato (±)-angustureina, a partir de um b-amino éster racêmico controlado por resolução cinética enzimática, é descrita. Esta estratégia permitiu, incorporar tanto o esqueleto básico como controlar o único estereocentro no carbono 2 de ambos enantiômeros. A sequência em cinco etapas a partir dos b-amino éster e o carboxilato de sódio quirais para a síntese de ambos os alcalóides foi feita com um rendimento global de 80 e 44%, respectivamente, e excelentes excessos enantiomericos (95 e 96%, respectivamente) e sem nenhuma proteção de grupos funcionais em todas as etapas.


Introduction
Galipea officinalis Hancock, a tree found mainly in the Northern region of South America, popularly known as "angostura", is one of twenty species in the genus Galipea Aublet.This plant is widely used in Venezuelan folk medicine, mainly for paralysis treatment of the motor system.It also provides a tonic used for the treatment of ailments such as dyspepsia, dysentery, chronic diarrhoea and fever. 1 A novel 2-substituted quinoline alkaloid was isolated from the bark of this plant by Jacquemond-Collet et al. 2 The same plant was later investigated by Rakotoson et al.,  3 who reported on the isolation of five quinoline alkaloids.On the other hand, reports have shown that angustureine, galipeine, cuspareine and galipinine exhibit anti-malarial and cytotoxic activities. 4It is believed that the medicinal properties of these species are related to the presence of various quinoline and tetrahydroquinoline alkaloids found in this plant. 3Due to their structural simplicities associated with the promising pharmacological activities, these alkaloids, especially angustureine, have attracted the attention of the synthetic organic community.For this alkaloid alone, nearly 21 different syntheses have already been described. 5 In our quest to synthesize natural products that possess remarkable pharmacological activities, we report here an alternative strategy for the enantioselective synthesis of both (R)-and (S)-angustureine 1 and 2, respectively, as a supplement to this team previously published work, 5  by applying enzymatic resolution to control the single stereocenter at C-2 of (R)-and (S)-angustureine 1 and 2.
To the best of our knowledge, this approach is the first work that leverages both the hydrolysis product and the corresponding ester for both stereoisomers in the synthesis of natural products.

Results and Discussion
Retrosynthetic analysis of (R)-and (S)-angustureine 1 and 2 The retrosynthetic analysis of (R)-and (S)-angustureine 1 and 2, outlined in Scheme 1, shows the key steps in the proposal.The source of the basic skeleton for both the (R)-and (S)-angustureine enantiomers comes from the racemic b-amino ester 6.The single stereocenter at carbon 2 of the chiral (S)-b-amino ester 5 and of the (R)-sodium carboxylate 7 can effectively be controlled from its racemate 6 by kinetic resolution using the protocol of Katayama et al. 6 The preparation of the iodides 4 and 12 from both (S)-b-amino ester 5 and (R)-sodium carboxylate 7, and their convergent coupling with allylmagnesium bromide 3, allowed the incorporation of the n-pentyl side chain at the 2-position to complete the enantioselective synthesis of both (R)-and (S)-angustureine enantiomers 1 and 2.
Preparation of the (S)-and (R)-b-amino esters 5 and 8 by enzymatic resolution The synthesis of (R)-and (S)-angustureine stereoisomers 1 and 2 begins with the preparation of racemic methyl 2- (1,2,3,4-tetrahydroquinolin-2-yl)acetate 6.This intermediate was prepared using the methodology described by Nagata et al. 7 The kinetic resolution of racemic ester 6 by treatment with Novozym ® 435 was performed in a 5% aqueous solution of tetrahydrofuran (THF) under stirring at 30 °C for 3 days.After filtration, the (S)-b-amino ester 5 (54% yield and 91% ee) was obtained by adding an aqueous NaHCO 3 solution to remove the (R)-sodium carboxylate 7, in addition to washing with water, extracting and purifying of the crude product by column chromatography (Scheme 2).
To compare retention times and check the enantiomeric ratio of both the (S)-and (R)-b-amino esters 5 and 8 from their respective chromatograms, the (R)-b-amino ester 8 was prepared.The aqueous layer containing the (R)-sodium carboxylate 7 (Scheme 2) from the enzymatic resolution reaction was dried under heating in vacuum.The ester 8 was prepared by treating the carboxylate 7, it was dissolved in methanol, with thionyl chloride, and refluxed.After the addition of saturated sodium bicarbonate, extraction and concentration, the residue was purified to give the (R)-b-amino ester 8 (30% yield and 96% ee).
Preparation of electrophiles 4, 9 and 10 and synthesis of (R)-angustureine 1 The synthesis of (R)-angustureine 1, first involved the preparation of the electrophilic substrates: iodide 4, mesylate 9 and bromide 10 from (S)-b-amino ester 5 in an attempt to optimize the convergent coupling with the allylmagnesium bromide 3. The iodide 4 was prepared in a one pot sequence in three steps.N-methylation of 5 was performed by treatment with iodomethane and K 2 CO 3 , followed by the reduction of the ester function with LiAlH 4 .Iodination of the resulting alcohol by treatment with I 2 and PPh 3 , produced the iodide 4 (89%, three steps), as shown in Scheme 3. The mesylate 9 was prepared following the same procedure above for the two first steps from 5, followed by treatment of the resulting alcohol with MsCl, in turn yielding the corresponding mesylate 9 (82% yield, three steps).Bromide 10 was prepared from mesylate 9 by refluxing with LiBr in THF for 24 h.
With the electrophiles 4, 9 and 10 in hand, attempts at convergent coupling with allylmagnesium bromide 3 for the synthesis of (R)-angustureine 1 were begun.These first attempts were carried out at -78 °C and at room temperature.Unfortunately, all of these attempts were unsuccessful since only traces of the products could be obtained when all these reactions were carried out at these temperatures.Due to these disappointing results, the parameter of temperature had to be changed.Further attempts to couple iodide 4 with Grignard 3 under reflux were performed, which surprisingly, generated a high yield of the desired crude product (Scheme 3).After having completed the hydrogenation of the double bond of the side chain of the resulting alkene, the residue was then purified by flash chromatography to give 1 (90% yield and 95% ee, two steps) as a pale yellow oil; [a] D −28.6 (c 1.0, CHCl 3 ) ([a] D −7.16 ( c 1.0, CHCl 3 )). 5  Preparation of iodide 12 and synthesis of (S)-angustureine 2Moreover, for the synthesis of (S)-angustureine 2, a change was needed to be made in the esterification step of the carboxylate 7. Thus, the yield of this step was very low when the (R)-b-amino ester 8 was prepared (see Scheme 2).A further analysis showed that, in addition to the esterification of carboxylate 7, the N-methylation may also be possible if carried out in a single step.Therefore, treatment of 7 with iodomethane and K 2 CO 3 in dimethylformamide (DMF) generated the N-methyl-b-amino ester 11 (52% yield), as can be seen in Scheme 4. The subsequent steps were carried out following the same procedure as described for the synthesis of (R)-angustureine 1 in Scheme 3. Reduction of the ester function and iodination of the resulting alcohol yielded iodide 12 (94% yield, two steps).Then, treatment of 12 with allylmagnesium bromide 3, and after hydrogenation of the double bond of the resulting alkene produced 2 (90% yield and 96% ee, two steps) as a pale yellow oil; [a] D +25.9 (c 1.0, CHCl 3 ), ([a] D +7.9 (c 1.0, CHCl 3 )). 5The spectra data of both (R)-and (S)-angustureine enantiomers 1 and 2 are fully consistent with those reported in the literature. 5However, a deviation could be observed as regards their specific rotations, [a] D +7.9 (c 1.0, CHCl 3 ) and [a] D −7.16 (c 1.0, CHCl 3 ) 5 for (S)-and (R)-enantiomers, which may well be related to the high enantiomeric purity of our samples, given that, in the present work, a high levo-and dextrorotatory correlation could be identified between both synthesized enantiomers, [a] D +25.9 (c 1.0, CHCl 3 ) and [a] D −28.6 (c 1.0, CHCl 3 ) for (S)-and (R)-angustureine, respectively.Scheme 3. Synthesis of (R)-angustureine 1.

Preparation of rac-angustureine 13
Finally, to compare retention times and determine the enantiomeric excesses of the (R)-and (S)-angustureine stereoisomers 1 and 2, from their respective chromatograms, rac-angustureine 13 was prepared (see Supplementary Information).Angustureine 13 was synthesized from rac-b-amino ester 6, in a similar manner as performed with chiral angustureine 1, shown in Scheme 3, but inverting the sequence of the three first steps (i.e., reduction, iodination and N-methylation to give rac-iodide 14) in an attempt to improve the yield.However, this change generated only an 80% yield of 13 (five steps), which was equal to the yield for 1.

Conclusions
In summary, the total synthesis of both (R)-and (S)-angustureine stereoisomers 1 and 2 was achieved in five steps, starting from chiral (S)-amino ester 5 and (R)-sodium carboxylate 7, with overall yields of 80 and 44%, respectively.The single stereocenter at C-2 of 5 and 7 was controlled by kinetic enzymatic resolution from rac-b-amino ester 6.This is the first work that leverages both the hydrolysis product and the corresponding ester for the enantiomeric synthesis of both natural products applying an enzymatic resolution.Further studies on the synthesis of other congeners applying this same methodology are currently in progress.