Letter pubs.acs.org/OrgLett
Combined Effect of the Picoloyl Protecting Group and Triflic Acid in Sialylation Samira Escopy, Scott A. Geringer, and Cristina De Meo* Department of Chemistry, Southern Illinois University Edwardsville, Edwardsville, Illinois 62026, United States S Supporting Information *
ABSTRACT: The stereoselective synthesis of sialosides is still one of the major challenges in carbohydrate chemistry. The synthesis and glycosidation of novel sialyl donors bearing a picoloyl substituent at C-4 are reported. High stereoselectivities and faster reactions were observed in the presence of an excess of triflic acid. The acid excess does not have the same effect on conventional sialyl donors, which suggests a possible synergistic effect of the picoloyl substituent and the triflic acid.
S
in we report our preliminary data on the effect of the O-picoloyl (Pico) group at C-4 in sialylations with primary acceptors. Reported for the glycosyl donors of the gluco-,22 galacto-,21 and manno-series,23 the HAD reaction provided high facial selectivities, always syn in respect to the remote Pico substituent, in glycosylation. The HAD reaction was particularly efficient under high-dilution conditions when DMTST or NIS/TfOH were used as promoters.24 High facial selectivity could also be achieved at standard concentrations (0.05 M) by the use of Br2 as the promoter.21 In order to investigate whether the Pico group could mediate the α-facial attack of the aglycone in sialylations, we synthesized 4-O-picoloylated sialyl donors 3a,b from their corresponding peracetylated counterparts 1a,b25,26 (Scheme 1). Somewhat lower stereoselectivity reported for phenylthio glycosyl donors in the HAD reaction21 prompted us to investigate different leaving groups, ethylthio in 3a and phenylthio in 3b. The synthesis of 4-O-picoloylated compounds 3a,b was accomplished by de-O-acetylation of peracetylated precursors
ialic acids and conjugates thereof are complex natural molecules that are involved in numerous biological phenomena, ranging from cell−cell adhesion and mobility to oncogenesis and recognition by viruses and bacteria.1,2 Sialylations by chemical methods are often complicated by the intrinsic structural features of sialic acids. Therefore, the stereoselective synthesis of sialosides remains one of the challenges of carbohydrate chemistry.3,4 Recently, significant attention has been given to a deeper understanding of the mechanism of glycosylation, and the influence of contributing factors such as counterions, solvent and protecting groups.5−7 In the sialic acid field, one of the major advances has been the discovery of how the C-5 effect can modulate the reactivity of sialyl donors and their stereoselectivity in sialylation.6,8−13 However, the effect of O-substitution of sialic acids beyond traditional acetates remains largely unexplored, whereas in the mainstream of carbohydrate chemistry the study of Oprotecting groups occupies a very important niche and represents a major venue of research. Clearly, O-protecting groups can have a crucial role in stabilizing intermediates, directing or hindering the nucleophilic attack of the acceptor, as well as influencing the dipole moment of the reactants.5,14 Following the pioneering research by Ye,15 Woerpel,16 and many other groups,17,18 our laboratory recently demonstrated that (1) during reactions via oxacarbenium ion intermediates a conformational change to the all-axial conformation can occur;19 (2) per-O-benzoylation of sialic acid creates a βdirecting sialyl donor, and (3) the effect of the tertbutyldimethylsilyl (TBDMS) group at C-4 is activating and α directing only in sialylations in acetonitrile as a solvent (MeCN).20 Thus, as a part of a general research goal toward a detailed investigation of the effect of O-protecting groups in sialylation reactions, and inspired by the data reported by Demchenko and co-workers on the development of the hydrogen-bond-mediated aglycone delivery (HAD),21−24 here© 2017 American Chemical Society
Scheme 1. Synthesis of Sialosides 3a,b
Received: March 31, 2017 Published: April 28, 2017 2638
DOI: 10.1021/acs.orglett.7b00976 Org. Lett. 2017, 19, 2638−2641
Letter
Organic Letters
conditions (0.005 M) that were found to be very beneficial for the HAD reactions with hexoses.24 However, the somewhat low reactivity profile of donors 3a,b, along with the intrinsic propensity of sialyl donors to eliminate, led to the observation that reactions at high dilution conditions (0.005 M) result in elimination rather than sialylation. As a result, the respective 2,3-dehydro derivative was found to be the predominant or even exclusive product of the reaction. A more attentive investigation of sialyl donor 3a using 1H NMR spectroscopy showed a concentration-dependent shift of the proton signals corresponding to H-4 and H-5, which led us to propose that an intermolecular H bond existed between the donors and that this was responsible for the poor HAD outcome. To investigate whether H-bonding with the 5-acetamido group was diminishing the HAD effect of the 4-Pico substituent, we turned our attention to investigating sialyl donors incapable of H-bonding between NH and the picoloyl group. For this purpose, we obtained novel sialyl donors 8a and 8b equipped with the 5-(N-acetyl)acetamido group, and as a comparison, the 5-(N-acetyl)acetamido sialyl donor 7 lacking the Pico group. We were also curious to investigate whether the 5-(N-acetyl)acetamido protection would enhance the reactivity of sialyl donors 8a and 8b, similar to that observed for similar sialyl donors.28 which would be beneficial for our study. The synthesis of di-N-acetylated sialyl donors 8a,b was accomplished from the corresponding monoacetylated counterparts 3a,b by the treatment with isopropenyl acetate in the presence of p-toluenesulfonic acid in DMF. As listed in Table 2, for the per-O-acetylated donor series the additional N-acetyl group in donor 7 provided a decreased
1a,b with NaOMe followed by the regioselective protection at C-8 and C-9 with dimethoxypropane (DMP). The regioselective introduction of the Pico group at C-4 was then performed using 2-picolinic acid in the presence of 1-ethyl-3(3-(dimethylamino)propyl)carbodiimide (EDC) as the coupling agent and DMAP to afford 2a,b in 87% and 82% yields, respectively (Scheme 1). Removal of the isopropylidene group in intermediates 2a,b and subsequent per-acetylation gave target compounds 3a,b in good yields (67% and 86%, respectively). It is noteworthy that no regioselection was observed for picoloylation in the presence of N,N′-dicyclohexylcarbodiimide (DCC) as the coupling reagent. This reaction led to the formation of the 4,7-di-O-picoloylated compound as the major product. Our investigation of the sialyl donor properties began with the coupling of the phenylthio sialyl donor 3a with galactosyl acceptor 4 using standard promoters in standard amounts for sialylation reaction, i.e., N-iodosuccinimide (NIS, 2.0 equiv/ equiv donor) and triflic acid (0.7 equiv/equiv donor). However, we made some essential adjustments to the reaction conditions as follows. Although acetonitrile is the most common solvent for sialylations,3,4 we feared that this participating solvent might interfere with the course of the HAD pathway. Therefore, our initial investigation was performed using dichloromethane as the solvent. Sialylation reactions in dichloromethane typically require a super low temperature of −78 °C to enhance stereoselectivity and suppress competing elimination. For example, sialyl donor 1a gave disaccharide 5 in 66% yield with a prevalence of the α-anomer (α/β = 1.5:1). However, no activation of donors 3a,b was observed at this temperature, which suggested a deactivating effect of the 4-O-Pico group. Thus, by gradually increasing the temperature, we observed that donors 3a,b could only be activated at −45 °C and above. To acquire a reliable comparison point, we performed test reactions with the benchmark sialyl donors 1a and 1b at this unusual temperature. Sialylations of acceptor 4 with donors 1a and 1b gave the desired target disaccharide 5 in 68% and 94% yield, respectively, albeit with no stereoselectivity (entries 1 and 3, Table 1). On the other hand, the C-4-picoloylated
Table 2. Sialylation with 5-(N-Acetyl)acetamido Donors
Table 1. Sialylations with Acetylated vs 4-O-Picoloylated Donors
entry
donor
time (min)
product
yield (%)
α/β ratio27
1 2 3 4
1a 3a 1b 3b
60 300 10 45
5 6 5 6
68 70 94 75
1.1:1 3.4:1 1.1:1 4.8:1
entry
donor
time (min)
product
yield (%)
α/β ratio
1 2 3
7 8a 8b
5 240 15
9 10 10
73 94 60
1:2.2 3.8:1 4.0:1
reaction time (5 min) and an improved yield (73%, entry 1) in comparison to that obtained with mono-N-acetylated donor 1a (1 h, 68% yield, Table 1). However, this reaction also led to a decrease in the stereoselectivity and disaccharide 9 was obtained with the prevalence of the β-anomer (α/β = 1:2.2). On the other hand, we were pleased to discover that the 5-(Nacetyl)acetamido protection offered a significant enhancement of sialylation with donors of the picoloylated series. Thus, di-Nacetylated 8a gave disaccharide 10 in a higher yield of 94%, along with a slightly enhanced stereoselectivity and reaction time, in comparison to those observed for its mono-Nacetylated counterpart 3a (entry 2, Table 2 vs entry 2, Table 1). A significant enhancement in reactivity was observed for the S-ethyl donor 8b in comparison to that observed for its monoN-acetylated counterpart 3a (entry 3, Table 2 vs entry 3, Table 1). However, the higher reactivity of the S-ethyl donor 8b was probably the culprit of lower sialylation yield and stereoselectivity observed for the synthesis of 10 (entry 3, Table 2). At this point, we decided to focus on the S-phenyl leaving
counterparts 3a and 3b gave disaccharide 6 in good yields (70% and 75%) and higher stereoselectivities (α/β = 3.4:1 and 4.8:1, respectively, entries 2 and 4, Table 1). As expected from our preliminary low-temperature experimentation, the reaction rates with 4-Pico sialyl donors 3a,b were much lower than those observed with their per-acetylated counterparts 1a,b. In order to enhance the stereoselectivity, we attempted to carry out our sialylation reactions under high dilution 2639
DOI: 10.1021/acs.orglett.7b00976 Org. Lett. 2017, 19, 2638−2641
Letter
Organic Letters group for its more balanced performance in terms of reactivity, stereoselectivity, and yield. Being committed to further enhancement of the reaction yields and stereoselectivities, we decided to explore additional reaction factors. After some exploratory experimentation with donor 3a, we noticed that an increase in the amount of triflic acid co-promoter gave a remarkable outcome. The result of this study is summarized in Table 3. Thus, increasing the amount of
Table 4. Sialylations of Sialyl Donor 3a with Acceptors 11 and 12
Table 3. Effect of Triflic Acid Molar Ratio on Sialylation of Acceptor 4 with Sialyl Donor 3a
entry
acceptor
1 2 3 4
11 11 12 12
TfOHa (time, h) 0.7 2.0 0.7 2.0
product (yield, %)
(16) (0.75) (16) (0.75)
13 13 14 14
(93) (70) (46) (51)
α/β ratio27 1.2:1 6.5:1 3.2:1 8.5:1
a
The amount of TfOH is provided in the number of equiv per 1 equiv of 3a.
entry
TfOHa
time (h)
yield of 6 (%)
α/β ratio27
1 2 3b 4 5b 6b
0.2 0.7 1.2 2.0 4.0 5.0
16 5 2 0.75 0.67 0.67
64 70 N/A 93 N/A N/A
3.0:1 3.0:1 3.0:1 14.2:1 9.0:1 8.0:1
To rationalize this data, we also investigated sialylation reactions at higher triflic acid molar ratio for per-O-acetylated donor 1a and the picoloylated donor 8a, with the additional acetyl group at N-5. During the attempted coupling of donor 1a with acceptor 4 in the presence of an excess of triflic acid, a faster donor consumption was observed, but no significant difference was observed in comparison with the reaction employing the standard amount of TfOH. A comparison of results listed in entry 1 in Table 1 and entry 1 in Table 5 shows
a
The amount of TfOH is provided in the number of equiv per 1 equiv of 3a. bThe anomeric ratios were determined by comparison of the integral intensities of the corresponding anomeric signals in the crude reaction mixture.
Table 5. Excess Triflic Acid Does Not Improve Sialylations with Donors 1a and 15 Lacking the Picoloyl Substituent
triflic acid from the commonly used substoichiometric amounts (0.2−0.7 equiv/equiv donor, entries 1 and 2) to an excess (2.0 equiv/equiv donor, entry 3 and beyond, entries 4 and 5) led to reduced reaction times for the glycosidation of donor 3a with acceptor 4. Strikingly, following the same trend the stereoselectivity also increased dramatically, peaking at 2.0 equiv (α/β = 14:1, entry 3). As shown in Table 3, adding up to 5.0 equiv of TfOH did not have a significant effect on the reaction rates but led to a somewhat declined stereoselectivity (entries 5 and 6). To our knowledge, neither a successful sialylation reaction nor the HAD reaction had been reported in the presence of an excess of triflic acid. In fact, a decrease in stereoselectivity was reported in HAD reactions performed in the presence of larger amounts of triflic acid.24 To elaborate upon this observation and to broaden the scope of the dramatic improvement in the outcome of the sialylation reaction in the presence of 2.0 equiv of triflic acid, we tested the coupling of donor 3a with other primary galactosyl acceptors 1129 and 12.30 The main results of this study are outlined in Table 4. Both highly reactive benzylated acceptor 11 and its less reactive benzoylated counterpart 12 gave comparable results. Enhanced reaction rates and stereoselectivities were recorded for the synthesis of disaccharides 13 and 14 in the presence of excess TfOH (entries 1−4). Thus, the 3-fold increase of the amount of TfOH translated in a nearly 3-fold improvement in stereoselectivity. The stereoselectivity of the formation of disaccharide 13 increased from unimpressive (α/β = 1.2:1, entry 1) to commendable (α/β = 6.5:1, entry 2). The reaction rate also improved dramatically from 16 h to 45 min, leading to a decline in the yield (entries 1 and 2). Similarly, the stereoselectivity of the formation of disaccharide 14 increased from a moderate (α/β = 3.2:1, entry 3) to very good (α/β = 8.5:1, entry 4). The reaction time also dropped dramatically from 16 h to 45 min, leading to a slightly improved yield (entries 3 and 4).
entry
donor
time (min)
product
yield (%)
α/β ratio
1 2 3
1a 8a 15
20 25 15
5 10 16
63 63 44
1.5:1 4.8:1 1.0:1
practically the same yield and stereoselectivity. Similarly, sialyl donor 8a reacted faster, but in this case, somewhat compromised yield and stereoselectivity were achieved (compare entry 2, Table 2 vs entry 2, Table 5). These data suggest a cooperative effect is occurring in the series of 4-O-picoloylated sialyl donors. The involvement of both the picoloyl group at C-4 and the acetamido group at C-5 seem to be crucial for providing the higher stereoselectivity observed for donor 3a. To support this statement, we investigated sialyl donor 15 bearing a benzoyl group at C-4. It showed practically no improvement in stereoselectivity when an excess of triflic acid was employed. An exemplary outcome of sialylation with donor 15 is shown in entry 3 (Table 5). The data reported herein can be rationalized as follows: the higher amount of triflic acid might be needed to eliminate the competition between the Pico group and other basic species in the reaction mixture, such as NIS. In this case, the overall faster donor consumption can be due in part to the higher production rate of iodonium ions. The higher yield of the sialylation product and the higher stereoselectivity are instead due to the presence of both the picoloyl group at C-4 and the acetamido at C-5. While the original HAD mechanism seems improbable 2640
DOI: 10.1021/acs.orglett.7b00976 Org. Lett. 2017, 19, 2638−2641
Organic Letters
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under these reaction conditions, a different type of H-bonding between the protonated picoloyl group and the acceptor cannot be completely excluded. We believe that the protonation of the picoloyl group at C-4 is capable of making substantial changes in the initial hydrogen bond net. As a result, the reaction might be proceeding via the intermediacy of a new type of the oxacarbenium ion that is capable of directing the nucleophilic attack of the acceptor. To investigate the viability of this hypothesis, we tested a phenylthio donor bearing a picoloyl substituent at C-7. As anticipated, while a higher amount of triflic acid led to a decreased reaction time, we did not notice any improvement of sialylation yield and stereoselectivity. This observation ultimately confirms the importance of the relative position of the Pico group to ensure higher stereoselectivity. In addition, this result hints at the importance of positioning of the Pico group on the pyranose ring rather than the side chain. In the case of the 4-Pico group, the direct placement of the Pico group on the ring translates to a stronger effect that the protonation may have on the conformation changes en route to the product formation. As a possible extension of this study, we are currently testing different solvent systems such as acetonitrile as well as expanding the scope of glycosyl acceptors. For the latter, preliminary experimentation on secondary acceptors shows a predominance of elimination reactions probably due to the lower reactivity of the nucleophiles. We are therefore in the process of tuning the reactions conditions to improve the overall outcome. In conclusion, higher stereoselectivities and yields have been observed in sialylation reactions in the presence of an unusually high molar ratio of triflic acid (2.0) to the donor. This effect seems to be directly linked to the presence of the picoloyl group at C-4 and the adjacent acetamido group at C-5. Additional investigation of this reaction and application to other systems and targets are currently ongoing in our laboratory.
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REFERENCES
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ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.orglett.7b00976.
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Letter
Experimental procedures; 1H and 13C NMR spectra for all new compounds (PDF)
AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. ORCID
Cristina De Meo: 0000-0001-9830-582X Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS
This work was supported by the National Science Foundation (NSF, CHE-RUI No. 1465003). Dr. Winter and Mr. Kramer (University of Missouri, St. Louis) are thanked for HRMS determinations. 2641
DOI: 10.1021/acs.orglett.7b00976 Org. Lett. 2017, 19, 2638−2641
