Next Article in Journal
Probing the Interfacial Behavior of Type IIIa Binary Mixtures Along the Three-Phase Line Employing Molecular Thermodynamics
Next Article in Special Issue
Strategy for Designing Selective Lysosomal Acid α-Glucosidase Inhibitors: Binding Orientation and Influence on Selectivity
Previous Article in Journal
Multitarget Anticancer Agents Based on Histone Deacetylase and Protein Kinase CK2 Inhibitors
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Molecules
Volume 25
Issue 7
10.3390/molecules25071498
molecules-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Synthesis of Pyrrolidine Monocyclic Analogues of Pochonicine and Its Stereoisomers: Pursuit of Simplified Structures and Potent β-N-Acetylhexosaminidase Inhibition

by
Xin Yan
1,2,
Yuna Shimadate
3,
Atsushi Kato
3,*,
Yi-Xian Li
1,2,*,
Yue-Mei Jia
1,2,
George W. J. Fleet
4,5 and
Chu-Yi Yu
1,2,5,*
1
Beijing National Laboratory for Molecular Science (BNLMS), CAS Key Laboratory of Molecular Recognition and Function, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, China
2
University of Chinese Academy of Sciences, Beijing 100049, China
3
Department of Hospital Pharmacy, University of Toyama, 2630 Sugitani, Toyama 930-0194, Japan
4
Chemistry Research Laboratory, Department of Chemistry, University of Oxford, Mansfield Road, Oxford OX13TA, UK
5
National Engineering Research Center for Carbohydrate Synthesis, Jiangxi Normal University, Nanchang 330022, China
*
Authors to whom correspondence should be addressed.
Molecules 2020, 25(7), 1498; https://doi.org/10.3390/molecules25071498
Submission received: 21 February 2020 / Revised: 18 March 2020 / Accepted: 23 March 2020 / Published: 25 March 2020
(This article belongs to the Special Issue Iminosugars: Beyond Glycosidase Inhibition)
Browse Figures
Review Reports Versions Notes

Abstract

:
Ten pairs of pyrrolidine analogues of pochonicine and its stereoisomers have been synthesized from four enantiomeric pairs of polyhydroxylated cyclic nitrones. Among the ten N-acetylamino pyrrolidine analogues, only compounds with 2,5-dideoxy-2,5-imino-d-mannitol (DMDP) and pochonicine (1) configurations showed potent inhibition of β-N-acetylhexosaminidases (β-HexNAcases); while 1-amino analogues lost almost all their inhibitions towards the tested enzymes. The assay results reveal the importance of the N-acetylamino group and the possible right configurations of pyrrolidine ring required for this type of inhibitors.

Graphical Abstract

1. Introduction

Since its isolation from fungus Pochonia suchlasporia var. suchlasporia TAMA 87 in 2009 [1], pochonicine (1) (Figure 1) has been an attractive synthetic target due to its potent and specific inhibition of β-N-acetylhexosaminidases (β-HexNAcases), including β-N-acetylglucosaminidases (β-GlcNAcases) and β-N-acetylgalactosaminidases (β-GalNAcases) [1,2]. β-HexNAcases are associated with many crucial biological processes [3]. In fungi and insects, the enzymes play important roles in the metabolism of the polysaccharide chitin [4,5]; in mammals, β-HexNAcases participate in the regulation of cell signalling and influence protein expression, degradation and trafficking [6]. In humans, many diseases including lysosomal storage disorders [7], type-II diabetes [6], insulin resistance [8] and Alzheimer’s disease [9,10,11] can be attributed to abnormality of β-HexNAcases. Therefore, study of β-HexNAcase inhibitors may provide alternate strategies for discovery of therapeutic drugs.
As one of the most potent β-HexNAcase inhibitors, pochonicine (1) possesses a polyhydroxylated pyrrolizidine skeleton, in contrast to the polyhydroxylated piperidine ring present in the other two naturally occurring iminosugars, nagstatin (2) [12] and siastatin B (3) (Figure 1) [13]. The N-acetylamino group is the common structural feature that distinguishes these natural products from the other iminosugars [14]. A diverse range of potent synthetic β-HexNAcase iminosugar inhibitors have been reported including pyrrolidines (4 [15,16,17], 5 [18,19], 6 [20,21] and 7 [22]), piperidines (8 [23,24], 9 [25] and 10 [26]), azepanes (11 [27,28,29]) and azetidines (12 [30] and 13 [31]); almost all of them contain an N-acetylamino group (Figure 2) [32].
The novel structure and inhibition properties of pochonicine (1) led to the rapid report of total syntheses [2], together with its enantiomer [33] and analogues [34]. However, syntheses of such highly substituted bicyclic pyrrolizidines with up to seven contiguous stereogenic centres are long and complex [35,36]. In contrast, construction of monocyclic pyrrolidine iminosugars with only four chiral centres are easier to accomplish. The pyrrolidine core of the corresponding iminosugar frequently appears to be the HexNAcase pharmacophore [37,38,39], as the pyrrolidine sections are responsible for mimicking the transition-state of enzyme reaction [40]. Among these monocyclic iminosugars, a series of pyrrolidines containing acetamide groups which are essential for their β-HexNAcase inhibitions were reported (Figure 2), among which considerable examples were accomplished before the isolation of pochonicine (1) [41,42]. Expectedly, quite a number of these five-membered acetamide derivatives were found to be potent β-HexNAcase inhibitors [16,21,25,26]. In continuation of our interests in structure-activity relationship study of iminosugars [39,43,44,45,46], in this work, 20 stereoisomeric pyrrolidine analogues of pochonicine (1) were synthesized and systematically assayed as glycosidase inhibitors, in order to look for novel molecules with simplified structure and remained potent inhibitory activities.

2. Results and Discussion

2.1. Synthesis of 1-N-Acetylamino-2,5-Imino-1,2,5-Trideoxy-l-Mannitol hydrochloride (A-10)

Synthesis of acetamide modified pyrrolidines generally rely on asymmetric synthesis from achiral starting materials or begin with carbohydrate precursors [32]. In the second strategy, carbohydrate-derived nitrones have significant advantages due to their ready chirality, availability and versatile chemistry [47,48]. In this work, polyhydroxylated nitrones A–H (Figure 3) were readily prepared from the enantiomers of xylose, arabinose, lyxose and ribose by literature methods [16,49,50,51].
The d-xylose-derived nitrone A is a convenient starting material for the synthesis of 1-amino-2,5-imino-1,2,5-trideoxy-l-mannitol hydrochloride (A-11) and its 1-N-acetylamino derivative (A-10) [16]. Nucleophilic additions with trimethylsilyl cyanide (TMSCN) [16] and nitromethane [52] were studied as agents for the introduction of aminomethyl substituents. Reaction of nitrone A with TMSCN at room temperature provide hydroxylamine A-2 in 96% yield as the sole product, and its C-2 configuration was determined as S-configuration through NOESY experiments since a strong interaction of H-2 and H-4 was observed; in contrast, the aza-Henry reaction gave a pair of inseparable epimers (A-2′a and A-2′b) in a 1:1 ratio. Since the corresponding reduction products were also difficult to purify, the attempt to introduce aminomethyl groups by aza-Henry reaction was not further investigated. The addition product A-2 was treated with Raney Ni/H2 in the presence of Boc2O, and then deprotected to afford diamine A-3 in good yield. Final hydrogenation of A-3 gave 1-amino-2,5-imino-1,2,5-trideoxy-l-mannitol dihydrochloride (A-11) in quantitative yield (Scheme 1).
Treatment of diamine A-3 with acetic anhydride provided compound A-4 in high yield, but the attempt to release the secondary amine selectively by hydrochloric acid was unsatisfactory, giving the target product A-5 in only 19% yield together with part of diamine A-3 recovered (Scheme 2). Selective reduction of the N-O bond of hydroxylamine A-2, can be achieved in the presence of SmI2 according to the reported method in moderate yield [53]. Since excessive amount of SmI2 was needed in above step, other reduction conditions were also tried. Though the typical Zn-Cu(OAc)2-AcOH system [16,54] led to complex mixtures, the modified condition with zinc replaced by iron [55] provided the target amine A-6 in excellent yield with no effect on the nitrile. Subsequent N-protection gave the carbamate A-7, which was then hydrogenated to convert the cyanide group to the primary amine. However, the reduction to amine A-8 proceeded in low yield (20–30%). Though the remaining two steps, N-acetylation and deprotection both can go smoothly to afford their corresponding products, the unsatisfactory reduction yield of compound A-7 seriously reduced the total yield of the whole route. Selective acetylation of the primary amine A-3 was also tried by strictly control of the usage of acetic anhydride and reaction time [15,16], but no target product was obtained. Unexpectedly, acetylation with acetic acid in dichloromethane provided the monoacetylated compound A-5 in 18% yield (improved to 24% yield when anhydrous MgSO4 was added). However, further attempts to improve the reaction were unsuccessful. In contrast, the mild acylation reagent N,N′,N″,N′″-tetraacetylglycoluril [56] showed excellent selectivity for primary amine acetylation, giving the monoacetylated A-5 in 92% yield when refluxing together in dichloromethane. Final hydrogenation of the intermediate then furnished the target product A-10 quantitatively.

2.2. Synthesis of 1-Amino and 1-N-Acetylamino Modified Pyrrolidine Stereoisomers

According to the strategy in Scheme 1 and Scheme 2, other 18 stereoisomeric pyrrolidine analogues of pochonicine (1) were synthesized from the corresponding nitrones B−H. Reaction of TMSCN with nitrones gave exclusively trans-addition products in high yields, with nitrone E and F as exceptions. Cyanation of nitrone E and F both afforded a pair of diastereomers with trans/cis ratios as 63:37 and 61:39, respectively. The configurations of the newly constructed chiral centres in compound D-2, F-2b and H-2 were unambiguously confirmed by X-ray crystallographic analysis (See Supplementary Materials). The structures of their corresponding enantiomers C-2, E-2b and G-2 can also be confirmed since their NMR data were indeed identical. Reduction of the resulting hydroxylamines by Raney Ni/H2 in the presence of Boc2O and subsequent deprotection afforded diamines B-3−H-3, which were then acetylated on the primary amine groups to give compounds B-5−H-5. Final hydrogenation of intermediates B-3−H-3 and B-5−H-5 provided the target products, i.e., 4·HCl, C-10−H-10 and B-11−H-11 (Table 1).

2.3. Glycosidase Inhibition

The synthesized 1-N-acetylamino and 1-amino pyrrolidine analogues were assayed against a range of enzymes, as shown in Table 2 and Table 3.
Compound D-10 which resembles the pyrrolidine ring of pochonicine (1) exhibited potent inhibition of β-GlcNAcases from various resources including bovine liver, HL60 and Jack bean (IC50 2.8 μM, 10 μM and 0.12 μM, respectively). While compound 4·HCl, the 1-deoxy-1-N-acetylamino derivative of 2,5-dideoxy-2,5-imino-d-mannitol (DMDP) [57], also behaved as similar potent inhibitor of bovine liver and Jack bean β-GlcNAcase (IC50 4.7 μM and 0.21 μM, respectively). Both the two compounds were found to potent inhibitors of HL60 β-GalNAcase (IC50 9.5 μM and 8.8 μM, respectively). However, comparing to the natural product pochonicine (1), the very powerful inhibitor of β-GlcNAcases and β-GalNAcases, both the two analogues showed significant decrease in the inhibition of two enzymes. Unexpectedly, pochonicine (1) did not exhibit any inhibition of α-GalNAcases, but compound D-10 showed moderate inhibition of chicken liver α-GalNAcase (IC50 65.3 μM). For other 1-N-acetylamino compounds tested in Table 2, part of the compounds are only moderate or weak inhibition of the tested β-HexNAcases, and the other completely lost their β-HexNAcase inhibitory activities.
As shown in Table 3, 1-amino pyrrolidine analogues failed to provide positive assay results. Most of them showed weak or no inhibition of all the glycosidases tested. As an exception, compound D-11 with the pyrrolidine ring of pochonicine (1) were found to be moderate inhibitor of α-mannosidase (IC50 54 μM) and β-GlcNAcase (IC50 99 μM) from Jack bean, and α-GalNAcase (IC50 44 μM) from chicken liver. The assay results further indicated the importance of acetamide groups for β-HexNAcase inhibitors. However, the more potent inhibition of compound D-11 of α-GalNAcase than compound D-10 may indicate the significant role of the scaffold in interaction with active sites of the enzyme instead of the 1-N-acetylamino group.
Though all the compounds in Table 2 and Table 3 can also be regarded as 1-N-acetylamino and 1-amino derivatives of their corresponding pyrrolidines (for example, compound 4·HCl is the 1-deoxy-1-N-acetylamino derivative of DMDP), they lost almost all their inhibition towards other tested glycosidases including glucosidase, galactosidase, mannosidase, α-l-fucosidase, trehalase, amyloglucosidase, α-l-rhamnosidase and β-glucuronidase, revealing the importance of C-1 hydroxyl groups in interaction with the corresponding enzymes.
Therefore, both configurations of the pyrrolidine ring and the 1-N-acetylamino group have significant influences on the inhibition of β-HexNAcases and α-GalNAcase. In detail, 1-N-acetylamino pyrrolidine analogues with the same configuration as DMDP and pochonicine (1) showed powerful inhibition of these enzymes, revealing the importance of the right configurations of A ring. Furthermore, the results indicate that pochonicine analogue with the A ring in DMDP configuration may also turn out to be potent inhibitors of the above enzymes. The structure-activity relationship reported in this work may be helpful in pursuing simplified pochonicine (1) analogues and more potent glycosidase inhibitors.

3. Materials and Methods

3.1. General Methods

All reagents were used as received without any further purification or prepared as described in the literature. TLC plates were visualized by ultraviolet light or by treatment with a spray of Pancaldi reagent ((NH4)6MoO4, Ce(SO4)2, H2SO4, H2O) or a 0.5% solution of KMnO4 in acetone. Column chromatography was performed on a flash column chromatography with silica gel (200–300 mesh, Inno-chem, Beijing, China). NMR spectra were measured in CDCl3 (with TMS as internal standard) or D2O (with H2O as internal standard) on a Bruker AV300, AV400 or AV500 magnetic resonance spectrometer (Bruker, Ettlingen, Germany) (1H NMR at 300 MHz, 400 MHz or 500 MHz, 13C NMR at 125 MHz). High-resolution mass spectra (HRMS) were performed on a Thermo Fisher Exactive Spectrometer (Thermo Fisher Scientific, Waltham, MA, USA). Polarimetry was determined using an Optical Activity AA-10R polarimeter with concentrations (c) given in gram per 100 mL. Infrared spectra were recorded as films on KBr plates on a Nicolet-6700 FT-IR spectrometer (Thermo Fisher Scientific, Waltham, MA, USA).

3.2. Material and Methods for the Enzyme Inhibition Assay

With rat intestinal maltase as an exception, other enzymes were purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). Brush border membranes prepared from rat small intestine according to the method of Kessler et al. [58] were assayed at pH 6.8 for rat intestinal maltase using maltose. The released d-glucose was determined colorimetrically using the Glucose CII-test Wako (Wako Pure Chemical Ind.; Osaka, Japan). Other glycosidase activities were determined using an appropriate p-nitrophenyl glycoside as substrate in a buffer solution at the optimal pH value of each enzyme. The reaction was stopped by adding 400 mM Na2CO3. The released p-nitrophenol was measured spectrometrically at 400 nm [59].

3.3. Chemistry

3.3.1. General Procedure for Synthesis of Hydroxylamines A-2, B-2, C-2, D-2, E-2a, E-2b, F-2a, F-2b, G-2 and H-2 with A-2 as an Example

To a solution of nitrone A (1.25 g, 3.00 mmol) in THF (5 mL) and methanol (25 mL) was added dropwise TMSCN (0.45 mL, 3.60 mmol) under Ar atmosphere at 0 °C. After stirring at room temperature for 6–8 h, TLC showed completion of the reaction. The resulting solution was concentrated in vacuo, and the residue was purified by flash column chromatography (silica gel, petroleum ether/EtOAc = 6/1) to give hydroxylamine A-2 (colourless syrup, 1.28 g, 96% yield). Data for (2S,3S,4S,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (A-2) (Ref. [16]): [α]D23 −9.5 (c 1.1 in CH2Cl2); νmax/cm−1: 3366 (s), 3088 (m), 3063 (m), 3030 (s), 2920 (s), 2867 (vs), 2237 (w), 1497 (s), 1454 (vs), 1362 (s), 1207 (m), 1100 (vs), 1028 (s), 738 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.36–7.20 (m, 15H), 6.52 (s, 1H, OH), 4.54–4.37 (m, 6H), 4.20 (d, J = 1.5 Hz, 1H, H-2), 4.13 (t, J = 2.1 Hz, 1H, H-3), 3.93 (dd, J = 6.5 and 2.2 Hz, 1H, H-4), 3.72 (dd, J = 10.5 and 3.3 Hz, 1H, H-6), 3.54 (dd, J = 10.5 and 4.0 Hz, 1H, H-6′), 3.28–3.25 (m, 1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.4, 137.3, 136.4, 128.7, 128.4, 128.3, 128.0, 127.97, 127.93, 127.88, 127.82, 115.7, 83.6 (C-3), 81.2 (C-4), 73.3 (PhCH2), 72.3 (PhCH2), 72.0 (PhCH2), 69.4 (C-5), 66.4 (C-6), 61.2 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1941.
Data for (2R,3R,4R,5R)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (B-2): Colourless syrup, 1.26 g, 94% yield from nitrone B (1.26 g, 3.02 mmol); [α]D22 +8.7 (c 1.0 in CH2Cl2); νmax/cm−1: 3359 (m), 3031 (m), 2868 (m), 2239 (w), 1496 (m), 1454 (s), 1362 (m), 1207 (m), 1097 (vs), 1027 (m), 737 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.36–7.20 (m, 15H), 6.46 (s, 1H, OH), 4.55–4.37 (m, 6H), 4.20 (d, d, J = 1.5 Hz, 1H, H-2), 4.14 (t, J = 2.1 Hz, 1H, H-3), 3.94 (dd, J = 6.5 and 2.2 Hz, 1H, H-4), 3.72 (dd, J = 10.5 and 3.3 Hz, 1H, H-6), 3.54 (dd, J = 10.5 and 4.0 Hz, 1H, H-6′), 3.28–3.26 (m, 1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.4, 137.3, 136.4, 128.7, 128.4, 128.3, 128.0, 127.96, 127.93, 127.87, 127.82, 115.7, 83.6 (C-3), 81.2 (C-4), 73.3 (PhCH2), 72.3 (PhCH2), 72.0 (PhCH2), 69.4 (C-5), 66.4 (C-6), 61.1 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1937.
Data for (2S,3S,4R,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (C-2) (Ref. [60]): Colourless syrup, 1.32 g, 96% yield from nitrone C (1.30 g, 3.12 mmol); [α]D23 +8.1 (c 1.5 in CH2Cl2); νmax/cm−1: 3306 (m), 3030 (m), 2925 (s), 2855 (s), 2251 (w), 1497 (m), 1454 (s), 1362 (m), 1209 (m), 1144 (s), 1102 (s), 1027 (s), 736 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm.): 7.34–7.23 (m, 15H), 6.72 (s, 1H, OH), 4.69–4.44 (m, 6H), 4.33 (t, J = 5.5 Hz, 1H, H-3), 4.28 (d, J = 5.6 Hz, 1H. H-2), 4.22 (t, J = 5.7 Hz, 1H, H-4), 3.72 (dd, J = 9.6 Hz and 6.9 Hz, 1H, H-6), 3.66 (dd, J = 9.3 Hz and 6.7 Hz, 1H, H-6′), 3.55–3.51 (m, 1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.79, 137.70, 136.7, 128.6, 128.45, 128.43, 128.2, 128.0, 127.9, 127.89, 127.81, 116.4, 80.2 (C-3), 75.8 (C-4), 73.8 (PhCH2), 73.4 (PhCH2), 73.1 (PhCH2), 68.9 (C-5), 67.4 (C-6), 60.4 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1939.
Data for (2R,3R,4S,5R)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (D-2) (Ref. [61]): Colourless syrup, 1.19 g, 92% yield from nitrone D (1.22 g, 2.93 mmol); [α]D23 −10.2 (c 1.0 in CH2Cl2); νmax/cm−1: 3328 (m), 3030 (m), 2925 (s), 2870 (s), 2251 (w), 1497 (m), 1454 (s), 1362 (m), 1209 (m), 1145 (s), 1102 (s), 1027 (s), 736 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.36–7.22 (m, 15H), 6.78 (s, 1H, OH), 4.69–4.43 (m, 6H), 4.33 (t, J = 5.5 Hz, 1H, H-3), 4.28 (d, J = 5.6 Hz, 1H. H-2), 4.22 (t, J = 5.7 Hz, 1H, H-4), 3.72 (dd, J = 9.6 Hz and 6.9 Hz, 1H, H-6), 3.65 (dd, J = 9.3 Hz and 6.7 Hz, 1H, H-6′), 3.55–3.51 (m, 1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.79, 137.71, 136.7, 128.6, 128.46, 128.43, 128.2, 128.0, 127.89, 127.81, 116.5, 80.2 (C-3), 75.8 (C-4), 73.8 (PhCH2), 73.4 (PhCH2), 73.1 (PhCH2), 69.0 (C-5), 67.4 (C-6), 60.4 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1938.
(2R,3R,4R,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (E-2a) and (2S,3R,4R,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (E-2b): 92% total yield from nitrone E (1.34 g, 3.21 mmol).
Data for E-2a: White solid, 821 mg, 58% yield; m.p. 117–119 °C; [α]D23 +7.8 (c 0.7 in CH2Cl2); νmax/cm−1: 3308 (m), 3028 (m), 2870 (m), 2245 (w), 1497 (m), 1453 (m), 1361 (m), 1144 (m), 1109 (s), 1043 (m), 1028 (m), 736 (vs), 693 (vs); 1HNMR (500 MHz, CDCl3) δ (ppm):7.37–7.23 (m,15H), 5.75 (s, 1H, OH), 4.56–4.45 (m, 6H), 4.15 (dd, J = 5.6 and 1.4 Hz, 1H, H-2), 4.00 (dd, J = 6.1 and 1.4 Hz, 1H, H-4), 3.83 (t, J = 9.1 Hz,1H), 3.73 (m, 2H, H-6), 3.39–3.35 (m,1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.8, 137.2, 136.4, 128.6, 128.5, 128.4, 128.3, 128.0, 127.91, 127.90, 127.83, 127.81, 118.35, 83.9 (C-3), 80.0 (C-4), 73.5 (PhCH2), 72.4 (PhCH2), 72.3 (PhCH2), 69.1 (C-6), 67.1 (C-5), 63.2 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1938.
Data for E-2b: White solid, 491 mg, 35% yield; m.p. 111–114 °C; [α]D23 −38.1 (c 1.0 in CH2Cl2); νmax/cm−1: 3359 (m), 3031 (m), 2868 (m), 2237 (w), 1496 (m), 1454 (m), 1362 (m), 1207 (m), 1097 (s), 1027 (m), 737 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm):7.35–7.21 (m, 15H), 5.98 (s, 1H, OH), 4.59–4.48 (m, 6H), 4.37 (d, J = 6.4 Hz, 1H, H-2), 4.18 (dd, J = 7.7 and 4.3 Hz, 1H, H-4), 4.07 (dd, J = 6.3 and 4.4 Hz, 1H, H-3), 3.80 (dd, J = 9.7 and 5.7 Hz, 1H, H-6), 3.68 (dd, J = 9.6 and 5.7 Hz, 1H, H-6′), 3.63−3.59 (m, 1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.7, 137.5, 136.6, 128.6, 128.4, 128.3, 128.2, 127.9, 127.8, 127.78, 127.73, 114.6, 81.1 (C-3), 80.8 (C-4), 73.5 (PhCH2), 73.0 (PhCH2), 72.9 (PhCH2), 67.4 (C-6), 66.7 (C-5), 60.6 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1939.
(2S,3S,4S,5R)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (F-2a) and (2R,3S,4S,5R)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (F-2b): 96% total yield from nitrone F (1.50 g, 3.60 mmol).
Data for F-2a: White solid, 985 mg, 59% yield; m.p. 118–119 °C; [α]D23 −5.5 (c 1.0 in CH2Cl2); νmax/cm−1: 3326 (m), 3028 (m), 2911 (m), 2870 (m), 2245 (w), 1497 (m), 1454 (s), 1241 (m), 1216 (m), 1144 (s), 1110 (vs), 736 (vs), 693 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm):7.37–7.23 (m,15H), 5.70 (s, 1H, OH), 4.56–4.45 (m, 6H), 4.15 (dd, J = 5.7 and 1.4 Hz, 1H, H-2), 4.00 (dd, J = 6.1 and 1.4 Hz, 1H, H-4), 3.83 (dd, J = 9.2 and 7.7Hz,1H, H-3), 3.75–3.71 (m, 2H, H-6), 3.39–3.35 (m,1H); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.8, 137.2, 136.4, 128.6, 128.5, 128.4, 128.3, 128.0, 127.91, 127.90, 127.83, 127.81, 118.3, 83.9 (C-3), 80.0 (C-4), 73.5 (PhCH2), 72.4 (PhCH2), 72.3 (PhCH2), 69.1 (C-6), 67.1 (C-5), 63.2 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1938.
Data for F-2b: White solid, 612 mg, 37% yield; m.p. 113–115 °C; [α]D23 +35.3 (c 1.0 in CH2Cl2); νmax/cm−1: 3359 (m), 3031 (m), 2868 (m), 2239 (w), 1496 (m), 1454 (m), 1362 (m), 1207 (s), 1097 (s), 1027 (m), 737 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm):7.35–7.21 (m, 15H), 6.04 (s, 1H, OH), 4.59 –4.47 (m, 6H), 4.37 (d, J = 6.3 Hz, 1H, H-2), 4.18 (dd, J = 7.7 and 4.3 Hz, 1H, H-4), 4.07 (dd, J = 6.3 and 4.4 Hz, 1H, H-3), 3.80 (dd, J = 9.7 and 5.7 Hz, 1H, H-6), 3.68 (dd, J = 9.6 and 5.7 Hz, 1H, H-6′), 3.63 −3.59 (m, 1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.7, 137.5, 136.6, 128.6, 128.4, 128.3, 128.2, 127.9, 127.8, 127.78, 127.74, 114.6, 81.1 (C-3), 80.8 (C-4), 73.5 (PhCH2), 73.0 (PhCH2), 72.9 (PhCH2), 67.4 (C-6), 66.7 (C-5), 60.6 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1939.
Data for (2R,3R,4S,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (G-2) (Ref. [61]): White solid, 1.39 g, 87% yield from nitrone G (1.51 g, 3.62 mmol); m.p. 106–109 °C; [α]D23 +0.6 (c 1.0 in CH2Cl2); νmax/cm−1: 3403 (m), 3029 (m), 2922 (m), 2871 (m), 2242 (w), 1497 (m), 1453 (m), 1352 (m), 1224 (m), 1145 (s), 1101 (s), 1027 (s), 742 (s), 694 (vs); 1HNMR (500 MHz, CDCl3) δ (ppm):7.35–7.24 (m,15H), 6.05 (s, 1H, OH), 4.70–4.43 (m, 6H), 4.10–4.05 (m, 2H, H-3, H-2), 3.83 (t, J = 6.1 Hz, 1H, H-4), 3.53–3.51 (m, 2H, H-6, H-6′), 3.31−3.30 (m,1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.6, 137.4, 136.7, 128.6, 128.48, 128.44, 128.2, 128.13, 128.11, 127.98, 127.90, 127.88, 127.86, 118.5, 77.3 (C-3), 75.2 (C-4), 73.3 ((PhCH2)), 72.8 (PhCH2), 72.2 (PhCH2), 72.0 (C-5), 68.0 (C-6), 61.8 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1936.
Data for (2S,3S,4R,5R)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-1-hydroxypyrrolidine (H-2): White solid, 1.45 g, 91% yield from nitrone H (1.50 g, 3.60 mmol); m.p. 109–111 °C; [α]D23 −1.4 (c 1.1 in CH2Cl2); νmax/cm−1: 3407 (m), 3029 (m), 2873 (m), 2242 (w), 1497 (m), 1453 (m), 1352 (m), 1225 (m), 1149 (s), 1102 (s), 1035 (s), 1028 (s), 743 (vs), 694 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.23 (m,15H), 6.11 (s, 1H, OH), 4.68–4.43 (m, 6H), 4.10–4.05 (m, 2H, H-3, H-2), 3.81 (t, J= 5.1 Hz, 1H, H-4), 3.53–3.48 (m, 2H, H-6, H-6′), 3.31–3.28 (m,1H, H-5); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.6, 137.4, 136.7, 128.6, 128.49, 128.45, 128.2, 128.13, 128.11, 127.98, 127.91, 127.8, 118.6, 77.3 (C-3), 75.2 (C-4), 73.3 (PhCH2), 72.8 (PhCH2), 72.2 (PhCH2), 72.0 (C-5), 68.0 (C-6), 61.8 (C-2); HRMS (ESI): calcd for C27H28O4N2Na+ [M + Na+] 467.1941, found 467.1933.

3.3.2. General Procedure for Synthesis of Hydroxylamines A-3, B-3, C-3, D-3, E-3a, E-3b, F-3a, F-3b, G-3 and H-3 with A-3 as an Example

Hydroxylamine A-2 (1.28 g, 2.88 mmol) was dissolved in methanol (10 mL), followed by Boc2O (1.46 mL, 6.30 mmol) and Raney Ni (1.0 g, 60%). The suspension was stirred under hydrogen atmosphere for 4 h when TLC showed completion of the reaction. Hydrogen was replaced by nitrogen, and catalyst was removed from the reaction mixture by filtration. The filtrate was concentrated in vacuo to afford the crude product as a colourless oil. The intermediate was dissolved in dichloromethane (10 mL) and cooled to 0 °C, trifluoroacetic acid (0.54 mL, 7.20 mmol) was added dropwise. After stirring at room temperature for 3 h, TLC showed completion of the reaction. The mixture was neutralized by aqueous NaHCO3 and extracted with dichloromethane (3 × 10 mL). The organic phases were combined, washed with brine and dried over MgSO4. After concentrated in vacuo, the crude product was purified by flash column chromatography (silica gel, dichloromethane/methanol = 25:1) to give diamine A-3 (colourless syrup, 1.05 g, 85% yield for two steps). Data for (2S,3S,4S,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (A-3): [α]D23 −16.3 (c 1.0 in CH2Cl2); νmax/cm−1: 3031 (m), 2923 (s), 2865 (s), 1682 (vs), 1453 (m), 1203 (vs), 1130 (vs), 1027 (m), 737 (s), 721 (s), 697 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.24 (m, 15H), 4.55–4.49 (m, 6H), 3.91 (t, J = 3.8 Hz, 1H), 3.76 (t, J = 3.8 Hz, 1H), 3.56−3.50 (m, 2H), 3.32 (q, J = 5.0 Hz, 1H), 3.15–3.12 (m, 1H), 2.79–2.71 (m, 2H), 1.77 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.2, 138.16, 138.15, 128.44, 128.41, 127.8, 127.78, 127.75, 127.71, 127.6, 87.3, 86.1, 73.2, 71.9, 71.7, 70.3, 64.1, 61.6, 44.3; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2478.
Data for (2R,3R,4R,5R)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (B-3): Colourless syrup, 380 mg, 88% yield from hydroxylamine B-2 (450 mg, 1.01 mmol); [α]D23 +11.8 (c 0.8 in CH2Cl2); νmax/cm−1: 3030 (m), 2866 (m), 1682 (vs), 1496 (m), 1453 (m), 1204 (vs), 1130 (vs), 1027 (m), 736 (s), 720 (s), 697 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.24 (m, 15H), 4.55–4.49 (m, 6H), 3.91 (t, J = 3.8 Hz, 1H), 3.76 (t, J = 3.8 Hz, 1H), 3.56–3.50 (m, 2H), 3.32 (q, J = 5.0 Hz, 1H), 3.16–3.12 (m, 1H), 2.79–2.71 (m, 2H), 1.76 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.2, 138.16, 138.15, 128.44, 128.41, 127.8, 127.78, 127.75, 127.71, 127.6, 87.3, 86.1, 73.2, 71.9, 71.7, 70.3, 64.1, 61.6, 44.3; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2478.
Data for (2S,3S,4R,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (C-3): Colourless syrup, 372 mg, 86% yield from hydroxylamine C-2 (450 mg, 1.01 mmol); [α]D22 +13.3 (c 1.0 in CH2Cl2); νmax/cm−1: 3328 (m), 3030 (m), 2864 (m), 1495 (m), 1453 (s), 1143 (s), 1094 (s), 1027 (m), 736 (s), 696 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.32–7.25 (m, 15H), 4.73–4.43 (m, 6H), 4.04 (t, J = 4.1 Hz, 1H), 3.70–3.61 (m, 3H), 3.42 (q, J = 6.6 Hz, 1H), 3.30 (q, J = 6.3 Hz, 1H), 2.77 (dd, J = 12.6 and 4.2 Hz, 1H), 2.59 (dd, J = 12.6 and 6.5 Hz, 1H), 1.50 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.6, 138.2, 138.1, 128.44, 128.41, 128.3, 127.8, 127.78, 127.76, 127.6, 127.5, 81.9, 78.0, 73.3, 73.2, 72.3, 69.7, 61.9, 59.0, 45.2; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2479.
Data for (2R,3R,4S,5R)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (D-3): Colourless syrup, 372 mg, 86% yield from hydroxylamine D-2 (450 mg, 1.01 mmol); [α]D23 −19.6 (c 1.2 in CH2Cl2); νmax/cm−1: 3292 (w), 3029 (m), 2859 (m), 1585 (m), 1495 (m), 1453 (s), 1143 (s), 1094 (s), 1027 (m), 734 (s), 696 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.32–7.25 (m, 15H), 4.73–4.43 (m, 6H), 4.04 (t, J = 4.1 Hz, 1H), 3.70–3.61 (m, 3H), 3.42 (q, J = 6.6 Hz, 1H), 3.30 (q, J = 6.3 Hz, 1H), 2.77 (dd, J = 12.6 and 4.2 Hz, 1H), 2.59 (dd, J = 12.6 and 4.3 Hz, 1H), 1.50 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.6, 138.2, 138.1, 128.44, 128.41, 128.3, 127.8, 127.78, 127.76, 127.6, 127.5, 81.9, 78.0, 73.3, 73.2, 72.3, 69.7, 61.9, 59.0, 45.2; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2478.
Data for (2R,3R,4R,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (E-3a): Colourless syrup, 379 mg, 88% yield from hydroxylamine E-2a (450 mg, 1.01 mmol); [α]D23 +12.1 (c 1.0 in CH2Cl2); νmax/cm−1: 3062 (w), 3029 (m), 2923 (s), 2860 (s), 1688 (m), 1496 (m), 1453 (s), 1201 (m), 1093 (vs), 735 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.35–7.24 (m, 15H), 4.57–4.44 (m, 6H), 3.92 (dd, J = 4.6 and 1.1 Hz, 1H), 3.73 (dd, J = 9.2 and 6.0 Hz, 1H), 3.65 (dd, J = 4.5 and 1.1 Hz,1H), 3.62 (dd, J = 9.2 and 6.8 Hz, 1H), 3.52 (q, J = 5.95 Hz, 1H), 3.12–3.08 (m,1H), 2.85 (dd,12.6 and 4.9 Hz,1H), 2.76 (dd, J = 12.6 and 7.1Hz, 1H), 1.61 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.3, 138.2, 138.0, 128.48, 128.41, 128.3, 127.8, 127.79, 127.75, 127.72, 127.69, 127.63, 127.60, 85.8, 82.9, 73.4, 71.67, 71.64, 69.7, 66.0, 60.6, 45.5; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2477.
Data for (2S,3R,4R,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (E-3b): Colourless syrup, 387 mg, 90% yield from hydroxylamine E-2b (450 mg, 1.01 mmol); [α]D23 −27.1 (c 0.9 in CH2Cl2); νmax/cm−1: 3062 (w), 3029 (m), 2920 (s), 2861 (s), 1495 (m), 1453 (s), 1362 (m), 1094 (vs), 1027 (m), 735 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.25 (m, 15H), 4.57–4.38 (m, 6H), 4.04 (dd, J = 4.6 and 2.3 Hz, 1H), 3.96 (dd, J = 5.3 and 2.3 Hz,1H), 3.67–3.60 (m, 2H), 3.55 (dd, J = 8.6 and 6.3 Hz,1H), 3.36 (q, J = 6.1 Hz,1H), 2.86–2.78 (m, 2H), 1.63 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.3, 138.2, 138.1, 128.4, 128.39, 128.35, 127.78, 127.74, 127.71, 127.6, 127.57, 127.56, 82.9, 82.5, 73.3, 72.2, 71.9, 69.5, 61.1, 58.5, 42.1; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2479.
Data for (2S,3S,4S,5R)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (F-3a): Colourless syrup, 359 mg, 83% yield from hydroxylamine F-2a (450 mg, 1.01 mmol); [α]D23 −9.3 (c 1.0 in CH2Cl2); νmax/cm−1: 3087 (w), 3029 (m), 2922 (s), 2859 (s), 1688 (m), 1496 (m), 1453 (s), 1363 (m), 1201 (s), 1094 (vs), 736 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm):7.35–7.24 (m, 15H), 4.58–4.44 (m, 6H), 3.92 (dd, J = 4.6 and 1.2 Hz, 1H), 3.73 (dd, J = 9.2 and 6.0 Hz, 1H), 3.65 (dd, J = 4.1 and 1.2 Hz,), 3.62 (dd, J = 9.2 and 6.8 Hz, 1H), 3.54–3.50 (m, 1H), 3.12–3.08 (m, 1H), 2.84 (dd, J = 12.6 and 4.9 Hz,1H), 2.76 (dd, J = 12.6 and 7.1Hz, 1H), 1.60 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.3, 138.2, 138.0, 128.48, 128.41, 128.3, 127.8, 127.79, 127.72, 127.69, 127.63, 127.60, 85.8, 82.9, 73.4, 71.69, 71.64, 69.7, 66.0, 60.6, 45.5; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2478.
Data for (2R,3S,4S,5R)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (F-3b): Colourless syrup, 367 mg, 85% yield from hydroxylamine F-2b (450 mg, 1.01 mmol); [α]D22 +23.8 (c 1.0 in CH2Cl2); νmax/cm−1: 3062 (w), 3029 (m), 2922 (s), 2858 (s), 1495 (m), 1453 (s), 1362 (m), 1093 (vs), 1027 (m), 734 (s), 696 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.25 (m, 15H), 4.57–4.38 (m, 6H), 4.04 (dd, J = 4.6 and 2.3 Hz, 1H), 3.96 (dd, J = 5.3 and 2.3 Hz, 1H), 3.67–3.60 (m, 2H), 3.55 (dd, J = 8.6 and 6.3 Hz, 1H), 3.36 (q, J = 6.1Hz, 1H), 2.86–2.78 (m, 2H), 1.62 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.3, 138.2, 138.1, 128.47, 128.42, 128.3, 127.8, 127.77, 127.74, 127.69, 127.60, 127.5, 82.9, 82.5, 73.4, 72.3, 71.9, 69.6, 61.1, 58.6, 42.1; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2478.
Data for (2R,3R,4S,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (G-3): Colourless syrup, 375 mg, 87% yield from hydroxylamine G-2 (450 mg, 1.01 mmol); [α]D22 +9.2 (c 1.1 in CH2Cl2); νmax/cm−1: 3362 (w), 3061 (m), 3029 (m), 2859 (s), 1496 (m), 1453 (s), 1363 (m), 1094 (vs), 1027 (m), 735 (vs), 697 (vs); 1H NMR (400 MHz, CDCl3) δ (ppm): 7.34–7.22 (m, 15H), 4.60–4.42 (m, 6H), 3.75 (t, J = 5.0 Hz, 1H), 3.55 (t, J = 5.9 Hz,1H), 3.49 (q, J = 5.0 Hz, 1H), 3.46–3.34 (m, 2H), 3.30 (dd, J = 10.8 and 5.9 Hz,1H), 2.78 (dd, J = 12.8 and 4.4 Hz, 1H), 2.60 (dd, J = 12.8 and 6.1 Hz,1H), 1.45 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3): δ (ppm): 138.31, 138.30, 138.27, 128.4, 128.3, 128.1, 127.9, 127.7, 127.69, 127.66, 79.4, 78.6, 73.2, 72.1, 71.7, 71.5, 62.9, 61.1, 44.9; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2479.
Data for (2S,3S,4R,5R)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine (H-3): Colourless syrup, 380 mg, 88% yield from hydroxylamine H-2 (450 mg, 1.01 mmol); [α]D23 −15.4 (c 1.0 in CH2Cl2); νmax/cm−1: 3061 (m), 3029 (m), 2861 (s), 1496 (m), 1453 (s), 1362 (m), 1122 (s), 1094 (s), 1027 (m), 736 (s), 697 (vs); 1H NMR (400 MHz, CDCl3) δ (ppm): 7.34–7.23 (m, 15H), 4.60–4.43 (m, 6H), 3.75 (t, J = 5.0 Hz, 1H), 3.55 (t, J = 5.9 Hz,1H), 3.49 (dd, J = 9.9 and 5.0 Hz, 1H), 3.46–3.34 (m, 2H), 3.30 (dd, J = 10.8 and 5.9 Hz,1H), 2.78 (dd, J = 12.8 and 4.4 Hz, 1H), 2.60 (dd, J = 12.8 and 6.1 Hz, 1H), 1.43 (s, 3H, NH, NH2); 13C NMR (125 MHz, CDCl3) δ (ppm): 138.3, 138.29, 138.26, 128.4, 128.3, 128.1, 127.9, 127.7, 127.69, 127.65, 79.4, 78.6, 73.2, 72.1, 71.7, 71.5, 62.9, 61.1, 44.9; HRMS (ESI): calcd for C27H33O3N2+ [M + H+] 433.2486, found 433.2479.

3.3.3. Synthesis of (2S,3S,4S,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-2-cyano-pyrrolidine (A-6)

To a suspension of iron powder (560 mg, 10.00 mmol, used as received) in acetic acid was added Copper (II) acetate (20 mg, 0.10 mmol), and the mixture was stirred at room temperature for 5–10 min until the bluish green suspension turned into reddish brown. The solution of hydroxylamine A-2 (450 mg, 1.01 mmol) in acetic acid (10 mL) was added, and the reaction mixture was stirred at room temperature overnight. Solvent was removed in vacuo, the residue was neutralized by aqueous NaHCO3 and filtered. The resulting filtrate was extracted with EtOAc (3 × 50 mL), then organic phases were combined, dried over MgSO4 and concentrated under reduced pressure. Purification by flash chromatography on silica gel (petroleum ether/EtOAc = 3/1) afforded pyrrolidine A-6 (light yellow syrup, 389 mg, 91% yield). Data for (2S,3S,4S,5S)-3,4-bis(benzyloxy)-5-(benzyloxymethyl) -2-cyano-pyrrolidine (A-6): [α]D23 −21.2 (c 1.0 in CH2Cl2); νmax/cm−1: 3293 (w), 3030 (w), 2926 (m), 2869 (m), 2246 (w), 1717 (m), 1661 (s), 1453 (s), 1397(m), 1149 (s), 1102 (s), 1027 (m), 736 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.36–7.25 (m, 15H), 4.59–4.45 (m, 6H), 4.27 (t, J = 3.1 Hz, 1H), 3.76 (d, J = 2.9 Hz, 1H), 3.83 (dd, J = 5.5 and 3.3 Hz, 1H), 3.59 (q, J = 6.5 Hz, 1H), 3.49–3.45 (m, 2H), 2.45 (br, 1H, NH); 13C NMR (125 MHz, CDCl3) δ (ppm): 137.7, 1377.5, 136.7, 128.6, 128.4, 128.2, 128.0, 127.87, 127.82, 127.78, 127.74, 119.3, 87.4, 84.1, 73.3, 72.5, 72.2, 69.7, 62.1, 51.9; HRMS (ESI): calcd for C27H29O3N2+ [M + H+] 429.2173, found 429.2181.

3.3.4. Synthesis of tert-butyl-(2S,3S,4S,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5-(benzyloxy methyl)pyrrolidine-1-carboxylate (A-8)

The mixture of compound A-6 (389 mg, 0.91 mmol) and Et3N (190 μL, 1.37 mmol) in dichloromethane (5 mL) was cooled by an ice-water bath, and Boc2O (298 mg, 1.37 mmol) was added. After stirring overnight at room temperature, the reaction was quenched by aqueous NaHCO3. The solution was then extracted with dichloromethane (3 × 10 mL), then organic phases were combined, dried over MgSO4 and concentrated in vacuo to give intermediate A-7 as a light yellow syrup (479 mg, 99% yield). The crude A-7 was directly dissolved in methanol (5 mL), and Raney Ni (500 mg, 60%) was added. The suspension was stirred under hydrogen atmosphere for 24 h when TLC showed part of intermediate A-7 remained unreacted. Longer reaction time did not lead to any further change. Hydrogen was then replaced by nitrogen, and catalyst was removed from the reaction mixture. The filtrate was concentrated in vacuo to afford a colourless oil, which was purified by flash chromatography (silica gel, dichloromethane/methanol = 50:1) to give compound A-8 (colourless syrup, 126 mg, 26% yield). Data for tert-butyl-(2S,3S,4S,5S)-2-(aminomethyl)-3,4-bis(benzyloxy)-5 -(benzyloxymethyl)pyrrolidine-1-carboxylate (A-8): [α]D23 −12.7 (c 0.6 in CH2Cl2); νmax/cm−1: 3029 (w), 2973 (m), 2929 (m), 1688 (s), 1453 (m), 1391 (vs), 1367 (s), 1160 (s), 1108 (s), 1027 (m), 734 (s), 697 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.37–7.23 (m, 15H), 4.77–4.41 (m, 6H), 4.18 (d, J = 7.7 Hz, 1H), 4.15–4.10 (m, 2H), 3.99–3.94 (m, 1H), 3.94–3.87 (m, 1H), 3.57 (d, J = 6.5 Hz,1H), 3.24 (d, J = 13.0 Hz, 1H), 2.87 (dd, J = 12.9 and 8.5 Hz, 1H), 1.39 (s, 9H); 13C NMR (125 MHz, CDCl3) δ (ppm): 156.7, 138.3, 137.8, 137.7, 128.38, 128.33, 128.2, 127.89, 127.84, 127.69, 127.60, 127.5, 127.4, 82.1, 80.3, 77.2, 73.2, 72.4, 72.1, 69.5, 60.3, 59.2, 44.4, 28.2; HRMS (ESI): calcd for C32H40O5N2+ [M + H+] 533.3010, found 533.3013.

3.3.5. Synthesis of tert-butyl-(2S,3S,4S,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl)pyrrolidine-1-carboxylate (A-9)

Compound A-8 (126 mg, 0.24 mmol) was dissolved in dichloromethane (5 mL), followed by Ac2O (28 μL, 0.29 mmol) and catalytic amount of DMAP. The solution was stirred at room temperature for 3-4 h, when TLC showed completion of the reaction. The solution was quenched by aqueous NaHCO3, and extracted with dichloromethane (3 × 10 mL). The organic phases were combined, dried over MgSO4 and concentrated in vacuo. The residue was purified by flash chromatography (silica gel, petroleum ether/EtOAc = 1/1) to afford compound A-9 (colourless syrup, 124 mg, 93% yield). Data for tert-butyl-(2S,3S,4S,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxy methyl)pyrrolidine-1-carboxylate (A-9): [α]D23 −19.1 (c 1.0 in CH2Cl2); νmax/cm−1: 3299 (w), 3029 (w), 2927 (m), 1690 (vs), 1682 (m), 1453 (m), 1390 (s), 1366 (s), 1274 (m), 1174 (m), 1096 (s), 1027 (m), 735 (m), 697 (m); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.33–7.25 (m, 15H), 6.64 (br, 1H, NHCO), 4.75–4.44 (m, 6H), 4.18 (dd, J = 8.8 and 6.1 Hz, 1H), 4.15–4.09 (m, 2H), 3.95–3.85 (m, 2H), 3.57 (d, J = 6.5 Hz,1H), 3.46–3.37 (m, 1H), 3.19–3.10 (m, 1H), 1.87 (s, 1H), 1.39 (s, 9H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.3, 155.5, 138.4, 138.1, 128.3, 128.2, 127.8, 127.7, 127.68, 127.62, 127.4, 80.7, 79.8, 77.8, 73.2, 72.5, 72.4, 70.0, 61.7, 58.6, 43.0, 28.3, 23.2; HRMS (ESI): calcd for C34H43O6N2+ [M + H+] 575.3116, found 575.3118.

3.3.6. General Procedure for Synthesis of Compounds A-5, B-5, C-5, D-5, E-5a, E-5b, F-5a, F-5b, G-5 and H-5 with A-5 as an Example

To the solution of compound A-3 (215 mg, 0.50 mmol) in dichloromethane (10 mL) was added N,N’,N’’,N’’’-tetraacetylglycoluril (170 mg, 0.55 mmol). The solution was refluxed for 3–5 h, when TLC showed disapearance of the starting material. The reaction was quenched by aqueous NaHCO3, and extracted with dichloromethane (3 × 10 mL). The organic phases were combined, dried over MgSO4 and concentrated under reduced pressure. The crude product was purified by flash chromatography (silica gel, dichloromethane/methanol = 50:1) to give compound A-5 (colourless syrup, 208 mg, 88% yield). Data for (2S,3S,4S,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5- (benzyloxymethyl)pyrrolidine (A-5): [α]D23 −7.7 (c 0.9 in CH2Cl2); νmax/cm−1: 3292 (w), 3030 (m), 2860 (m), 1656 (s), 1453 (m), 1363 (m), 1094 (s), 1027 (m), 737 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.25 (m, 15H), 6.08 (br, 1H, NHCO), 4.55–4.49 (m, 6H), 3.88 (t, J = 3.5 Hz, 1H), 3.76 (t, J = 3.4 Hz, 1H), 3.54–3.48 (m, 2H), 3.43–3.38 (m, 1H), 3.37–3.31 (m, 2H), 3.28–3.23 (m, 1H), 2.1 (br, 1H, NH), 1.88 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.2, 137.9, 137.83, 137.81, 128.46, 128.44, 127.87, 127.84, 127.81, 127.77, 127.73, 86.8, 85.5, 73.2, 72.0, 71.8, 69.8, 61.9, 61.0, 41.4, 23.2; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2588.
Data for (2R,3R,4R,5R)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (B-5) (Ref. [62]): Colourless syrup, 216 mg, 91% yield from diamine B-3 (215 mg, 0.50 mmol); [α]D23 +11.9 (c 1.0 in CH2Cl2); νmax/cm−1: 3283 (w), 3030 (m), 2859 (m), 1656 (s), 1454 (m), 1363 (m), 1093 (s), 1028 (m), 737 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.25 (m, 15H), 6.08 (br, 1H, NHCO), 4.55–4.49 (m, 6H), 3.88 (t, J = 3.5 Hz, 1H), 3.76 (t, J = 3.4 Hz, 1H), 3.54–3.48 (m, 2H), 3.43–3.38 (m, 1H), 3.37–3.31 (m, 2H), 3.28–3.23 (m, 1H), 2.16 (br, 1H, NH), 1.88 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.2, 137.9, 137.83, 137.81, 128.46, 128.44, 127.87, 127.84, 127.81, 127.77, 127.73, 86.8, 85.5, 73.2, 72.0, 71.8, 69.8, 61.9, 61.0, 41.4, 23.2; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2586.
Data for (2S,3S,4R,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (C-5): Colourless syrup, 201 mg, 85% yield from diamine C-3 (215 mg, 0.50 mmol); [α]D23 +11.9 (c 1.0 in CH2Cl2); νmax/cm−1: 3288 (w), 3029 (w), 2924 (m), 2855 (m), 1652 (s), 1453 (m), 1366 (m), 1089 (s), 1027 (m), 736 (s), 697 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.35–7.25 (m, 15H), 5.87 (br, 1H, NH), 4.73–4.49 (m, 6H), 4.00 (t, J = 4.0 Hz, 1H), 3.68−3.62 (m, 2H), 3.58 (t, J = 7.5 Hz, 1H), 3.44–3.38 (m, 2H), 3.35–3.25 (m, 2H), 2.16 (br, 2H, NH2), 1.89 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.3, 138.3, 138.0, 137.8, 128.47, 128.42, 128.3, 127.89, 127.88, 127.86, 127.80, 127.7, 127.6, 82.4, 77.6, 73.4, 73.2, 72.5, 69.5, 59.07, 59.04, 42.3, 23.2; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2588.
Data for (2R,3R,4S,5R)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (D-5): Colourless syrup, 194 mg, 82% yield from diamine D-3 (215 mg, 0.50 mmol); [α]D23 −2.5 (c 1.0 in CH2Cl2); νmax/cm−1: 3296 (w), 3028 (w), 2921 (m), 2855 (m), 1652 (s), 1554 (m), 1453 (s), 1365(m), 1091 (s), 1027 (m), 736 (s), 696 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.35–7.25 (m, 15H), 5.87 (br, 1H, NHCO), 4.73–4.49 (m, 6H), 4.00 (t, J = 4.1 Hz, 1H), 3.68–3.62 (m, 2H), 3.58 (t, J = 7.5 Hz, 1H), 3.44–3.38 (m, 2H), 3.35–3.25 (m, 2H), 2.16 (br, 1H, NH), 1.89 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.3, 138.4, 138.0, 137.9, 128.49, 128.44, 128.3, 127.9, 127.8, 127.7, 127.6, 82.4, 77.7, 73.4, 73.3, 72.6, 69.5, 59.09, 59.06, 42.3, 23.2; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2588.
Data for (2R,3R,4R,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (E-5a): Colourless syrup, 213 mg, 90% yield from diamine E-3a (215 mg, 0.50 mmol); [α]D23 +9.2 (c 1.0 in CH2Cl2); νmax/cm−1: 3295 (w), 3030 (w), 2924 (m), 2855 (m), 1652 (s), 1554 (m), 1493 (s), 1365 (m), 1091 (s), 1027 (m), 736 (s), 696 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.24 (m, 15H), 6.81 (br, 1H, NHCO), 4.58–4.42 (m, 6H), 4.19 (br, 1H, NH), 3.90 (d, J = 3.9 Hz, 1H), 3.77 (s, 1H), 3.73 (dd, J = 9.3 and 5.8 Hz, 1H), 3.68 (dd, J = 9.5 and 6.7 Hz, 2H), 3.62 (dd, J = 10.3 and 6.0 Hz, 1H), 3.48–3.43 (m, 1H), 3.39–3.34 (m, 1H), 1.76 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.6, 138.1, 137.7, 137.6, 128.54, 128.52, 128.4, 127.99, 127.91, 127.8, 127.79, 127.75, 127.6, 85.0, 82.2, 73.5, 71.9, 71.8, 69.6, 62.0, 60.4, 42.4, 22.8; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2585.
Data for (2S,3R,4R,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (E-5b): Colourless syrup, 194 mg, 82% yield from diamine E-3b (215 mg, 0.50 mmol); [α]D23 −17.2 (c 1.2 in CH2Cl2); νmax/cm−1: 3290 (m), 3063 (m), 3030 (m), 2925 (s), 2861 (s), 1651 (vs), 1549 (m), 1453 (s), 1367 (s), 1092 (vs), 1027 (m), 736 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.36–7.25 (m, 15H), 6.11 (br, 1H, NHCO), 4.56–4.35 (m, 6H), 3.99 (d, J = 1.6 Hz, 1H), 3.93 (d, J = 3.1 Hz, 1H), 3.63–3.61 (m, 2H), 3.57–3.52 (m, 3H), 3.30–3.25 (m, 1H), 2.36 (br, 1H, NH), 1.85 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.4, 138.1, 137.9, 137.7, 128.6, 128.4, 128.0, 127.89, 127.86, 127.84, 127.7, 82.6, 82.0, 73.4, 72.3, 72.1, 68.8, 58.9, 58.2, 39.6, 23.2; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2584.
Data for (2S,3S,4S,5R)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (F-5a): Colourless syrup, 206 mg, 87% yield from diamine F-3a (215 mg, 0.50 mmol); [α]D22 −4.1 (c 1.0 in CH2Cl2); νmax/cm−1: 3295 (w), 3064 (m), 3030 (m), 2924 (m), 2855 (m), 1652 (s), 1554 (m), 1493 (m), 1365 (m), 1091 (s), 1027 (m), 736 (s), 696 (s); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.23 (m, 15H) 6.54 (br, 1H, NHCO), 4.58–4.42 (m, 6H), 3.89 (dd, J = 4.1 and 0.6 Hz, 1H), 3.74–3.71 (m, 2H), 3.65 (dd, J = 8.9 and 6.9 Hz, 1H), 3.59 (dd, J = 10.0 and 5.4 Hz, 1H), 3.44–3.40 (m, 2H), 3.35–3.31 (m, 1H), 3.19 (br, 1H, NH), 1.73 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.6, 138.1, 137.7, 137.6, 128.54, 128.52, 128.4, 127.99, 127.9, 127.8, 127.79, 127.75, 127.6, 85.0, 82.2, 73.5, 71.9, 71.8, 69.6, 62.0, 60.4, 42.4, 22.8; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2587.
Data for (2R,3S,4S,5R)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (F-5b): Colourless syrup, 197 mg, 83% yield from diamine F-3b (215 mg, 0.50 mmol); [α]D22 +20.5 (c 1.0 in CH2Cl2); νmax/cm−1: 3293 (m), 3063 (m), 3030 (m), 2924 (s), 2861 (s), 1651 (vs), 1549 (m), 1453 (s), 1367 (s), 1092 (vs), 1027 (m), 736 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.35–7.24 (m, 15H) 6.24 (br, 1H, NHCO), 4.55–4.35 (m, 6H), 3.98 (dd, J = 6.0 and 1.9 Hz, 1H), 3.92 (dd, J = 4.9 and 1.9 Hz, 1H), 3.63 (m, 2H), 3.57–3.52 (m, 3H), 3.29–3.23 (m, 1H), 3.12 (br, 1H, NH), 1.84 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.4, 138.1, 137.9, 137.7, 128.6, 128.48, 128.42, 128.0, 127.89, 127.86, 127.85, 127.7, 82.5, 82.0, 73.4, 72.3, 72.1, 68.9, 58.9, 58.2, 39.6, 23.2; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2585.
Data for (2R,3R,4S,5S)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (G-5): Colourless syrup, 204 mg, 86% yield from diamine G-3 (215 mg, 0.50 mmol); [α]D23 +7.5 (c 1.0 in CH2Cl2); νmax/cm−1: 3293 (m), 3062 (m), 3029 (m), 2924 (s), 2861 (s), 1653 (vs), 1539 (m), 1453 (s), 1365 (m), 1100 (vs), 1027 (m), 736 (s), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.34–7.24 (m, 15H), 6.12 (br, 1H, NHCO), 4.58–4.45 (m, 6H), 3.77 (t, J = 4.7 Hz, 1H), 3.60 (t, J = 5.5 Hz, 1H), 3.50 (q, J = 4.4 Hz, 1H), 3.47–3.38 (m, 4H), 3.18–3.13 (m, 1H), 2.14 (br, 1H, NH), 1.74 (s, 3H); 13C NMR (125 MHz, CDCl3) δ (ppm): 170.2, 138.0, 137.9, 128.47, 128.40, 128.1, 128.0, 127.86, 127.80, 127.7, 80.0, 78.2, 73.3, 72.0, 71.6, 61.0, 59.6, 42.3, 23.0; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2587.
Data for (2S,3S,4R,5R)-2-N-acetylaminomethyl-3,4-bis(benzyloxy)-5-(benzyloxymethyl) pyrrolidine (H-5): Colourless syrup, 192 mg, 81% yield from diamine H-3 (215 mg, 0.50 mmol); [α]D23 −1.6 (c 1.2 in CH2Cl2); νmax/cm−1: 3304 (m), 3062 (m), 3030 (m), 2865 (s), 1651 (vs), 1549 (m), 1453 (s), 1365 (m), 1099 (vs), 1027 (m), 736 (vs), 697 (vs); 1H NMR (500 MHz, CDCl3) δ (ppm): 7.33–7.24 (m, 15H), 6.11 (br, 1H, NHCO), 4.57–4.43 (m, 6H), 3.76 (t, J = 4.8 Hz, 1H), 3.59 (t, J = 5.6 Hz, 1H), 3.49 (dd, J = 8.9 and 4.4 Hz, 1H), 3.47–3.37 (m, 4H), 3.18–3.13 (m, 1H), 2.14 (br, 1H, NH), 1.74 (s, 3H); 13C NMR (125 MHz, CDCl3): δ (ppm): 170.2, 138.0, 137.9, 128.47, 128.40, 128.1, 128.0, 127.86, 127.80, 127.7, 80.0, 78.2, 73.3, 72.0, 71.6, 61.0, 59.6, 42.3, 23.0; HRMS (ESI): calcd for C29H35O4N2+ [M + H+] 475.2591, found 475.2588.

3.3.7. General Procedure for Synthesis of 1-N-Acetylamino Derivatives (A-10, 4·HCl, C-10, D-10, E-10a, E-10b, F-10a, F-10b, G-10 and H-10) and 1-Amino Derivatives (A-11, B-11, C-11, D-11, E-11a, E-11b, F-11a, F-11b, G-11 and H-11) with A-10 as an Example

To a stirred solution of A-5 (95 mg, 0.20 mmol) and 3 N HCl (0.5 mL) in MeOH (10 mL) was added Pd/C (10 wt%, 30 mg) under Ar atmosphere and the reaction mixture was stirred under H2 atmosphere for 8 h. Then the catalyst was filtered and the solvent was removed under reduced pressure to afford compound A-10 (colourless syrup, 47 mg, 99% yield). Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-l-mannitol hydrochloride (A-10) (Ref. [16], reported as free base): [α]D23 −30.4 (c 0.5 in MeOH); νmax/cm−1: 3290 (vs), 2932 (s), 1646 (m), 1550 (m), 1369 (m), 1132 (m), 1041 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.25 (s, 1H,), 4.11 (s, 1H), 3.98 (dd, J = 12.0 Hz and 4.6 Hz, 1H), 3.95–3.91 (m, 1H), 3.86 (dd, J = 11.8 Hz and 8.7 Hz, 1H), 3.66 (d, J = 6.8 Hz, 2H), 3.63–3.61 (m, 1H), 2.04 (s, 3H, COCH3); 13C NMR (125 MHz, D2O): δ (ppm): 175.2, 75.7, 74.6, 67.5, 61.5, 59.2, 35.8, 21.7; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1181.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-d-mannitol hydrochloride (4·HCl) (Ref. [16,62], reported as free base): Colourless syrup, 50 mg, 99% yield from compound B-5 (101 mg, 0.21 mmol); [α]D25 +34.6 (c 0.5 in MeOH); νmax/cm−1: 3297 (vs), 2937 (s), 1644 (m), 1550 (m), 1369 (m), 1132 (m), 1042 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.22 (d, J = 2.0 Hz, 1H), 4.09 (t, J = 1.6 Hz, 1H), 3.95 (dd, J = 12.1 Hz and 4.8 Hz, 1H), 3.91 (dt, J = 6.9 Hz and 3.7 Hz, 1H), 3.83 (dd, J = 12.1 Hz and 8.6 Hz, 1H), 3.66 (d, J = 6.8 Hz, 2H), 3.61–3.58 (m, 1H), 2.01 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm): 175.2, 75.7, 74.6, 67.5, 61.5, 59.2, 35.8, 21.7; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1183.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-l-altritol hydrochloride (C-10) (Ref. [17], reported as free base): Colourless syrup, 45 mg, 99% yield from compound C-5 (90 mg, 0.19 mmol); [α]D22 −18.0 (c 0.7 in MeOH); νmax/cm−1: 3291 (vs), 2936 (s), 1635 (m), 1550 (m), 1419 (m),1369 (m), 1136 (m), 1042 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.33(s, 1H), 4.23 (dd, J = 9.3 Hz and 3.7 Hz, 1H), 3.99 (dd, J = 12.0 Hz and 4.8 Hz, 1H), 3.90 (dd, J = 11.6 Hz and 8.4 Hz, 1H), 3.82–3.80 (m, 1H), 3.72–3.66 (m, 2H), 3.59 (dd, J = 15.3 Hz and 7.7 Hz, 1H), 2.04 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm): 176.0, 72.5, 69.8, 62.0, 60.5, 57.4, 38.9, 21.6; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1186.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-d-altritol hydrochloride (D-10) (Ref. [16,61], reported as free base): Colourless syrup, 41 mg, 99% yield from compound D-5 (82 mg, 0.17 mmol); [α]D22 +21.3 (c 0.4 in MeOH); νmax/cm−1: 3296 (vs), 2936 (s), 1635 (m), 1550 (m), 1419 (m),1371 (m), 1136 (m), 1042 (m); 1H NMR (500 MHz, D2O) δ (ppm) 4.33 (t, J = 3.2 Hz, 1H), 4.23 (dd, J = 9.3 Hz and 3.7 Hz, 1H), 3.99 (dd, J = 12.0 Hz and 4.8 Hz, 1H), 3.90 (dd, J = 11.6Hz and 8.4Hz, 1H), 3.82–3.80 (m, 1H), 3.72–3.66 (m, 2H), 3.59 (dd, J = 15.3 Hz and 7.75 Hz, 1H), 2.04 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm): 176.0, 72.5, 69.8, 62.0, 60.5, 57.4, 38.9, 21.6; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1185.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-d-glucitol hydrochloride (E-10a) (Ref. [63], reported as free base): Colourless syrup, 46 mg, 99% yield from compound E-5a (91 mg, 0.19 mmol); [α]D25 +36.9 (c 0.85 in MeOH); νmax/cm−1: 3292 (vs), 2935 (s), 1645 (m), 1550 (m), 1369 (m), 1132 (m), 1045 (m); 1H NMR (400 MHz, D2O) δ (ppm): 4.27 (s, 1H), 4.12 (s, 1H), 3.98 (d, J = 9.4 Hz, 1H), 3.94–3.88 (m, 2H), 3.68–3.57 (m, 3H), 2.01 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm):175.8, 76.8, 74.5, 65.1, 63.0, 57.0, 39.4, 21.6; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1182.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-l-iditol hydrochloride (E-10b) (Ref. [64], reported as free base): Colourless syrup, 52 mg, 99% yield from compound E-5b (104 mg, 0.22 mmol); [α]D25 −9.6 (c 1.2 in MeOH); νmax/cm−1: 3292 (vs), 2936 (s), 1651 (m), 1553 (m), 1370 (m), 1136 (m), 1042 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.35 (d, J = 3.3 Hz, 1H), 4.28 (d, J = 2.0 Hz, 1H), 4.01–3.95 (m, 3H), 3.87 (dd, J = 11.9 and 8.3 Hz, 1H), 3.60 (d, J = 6.8 Hz, 2H), 2.01 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm): 175.8, 75.5, 74.1, 62.6, 61.3, 57.9, 38.6, 21.6; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1180.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-l-glucitol hydrochloride (F-10a): Colourless syrup, 44 mg, 99% yield from compound F-5a (88 mg, 0.19 mmol); [α]D25 −29.2 (c 1.05 in MeOH); νmax/cm−1: 3285 (vs), 2931 (s), 1645 (m), 1551 (m), 1370 (m), 1090 (m), 1063 (m); 1H NMR (400 MHz, D2O) δ (ppm): 4.30 (t, J = 1.6 Hz, 1H), 4.16 (t, J = 1.6 Hz, 1H), 4.01 (dd, J = 8.8 and 2.8 Hz, 1H), 3.95–3.88 (m, 2H), 3.69–3.59 (m, 3H), 2.04 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm):175.8, 76.8, 74.6, 65.2, 63.1, 57.0, 39.4, 21.6; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1183.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-d-iditol hydrochloride (F-10b): Colourless syrup, 42 mg, 99% yield from compound F-5b (83 mg, 0.17 mmol); [α]D25 +7.7 (c 1.0 in MeOH); νmax/cm−1: 3296 (vs), 2936 (s), 1651 (m), 1553 (m), 1371 (m), 1136 (m), 1041 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.34 (d, J = 3.4 Hz, 1H), 4.27 (t, J = 1.8 Hz, 1H), 4.00–3.94 (m, 3H), 3.86 (dd, J = 11.6 and 8.3 Hz, 1H), 3.60 (d, J = 6.9 Hz, 2H), 2.01 (s, 3H, COCH3); 13C NMR (125 MHz, D2O) δ (ppm): 175.8, 75.5, 74.1, 62.6, 61.3, 57.9, 38.6, 21.7; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1183.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-d-allitol hydrochloride (G-10): Colourless syrup, 46 mg, 99% yield from compound G-5 (92 mg, 0.19 mmol); [α]D23 −23.1 (c 1.0 in MeOH); νmax/cm−1: 3306 (vs), 2928 (s), 1635 (s), 1551 (s), 1419 (m),1370 (m), 1089 (m), 1063 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.22–4.19 (m, 2H), 3.93 (dd, J = 12.6 Hz and 3.8 Hz, 1H), 3.83 (dd, J = 12.6 Hz and 3.8 Hz, 1H), 3.78–3.74 (m, 1H), 3.73–3.66 (m, 2H), 3.57 (dd, J = 15.1 Hz and 7.4 Hz, 1H), 2.04 (s, 1H, COCH3); 13C NMR (125 MHz, D2O): δ(ppm): 176.1, 71.1, 70.2, 64.2, 62.8, 57.9, 38.5, 21.6; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1182.
Data for 1-N-acetylamino-2,5-imino-1,2,5-trideoxy-l-allitol hydrochloride (H-10): Colourless syrup, 41 mg, 99% yield from compound H-5 (81 mg, 0.17 mmol); [α]D23 +27.3 (c 0.95 in MeOH); νmax/cm−1: 3296 (vs), 2936 (s), 1651 (s), 1546 (s), 1419 (m), 1372 (m), 1091 (m), 1061 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.22–4.18 (m, 2H), 3.93 (dd, J = 12.5 Hz and 3.6 Hz, 1H), 3.82 (dd, J = 12.6 Hz and 3.6 Hz, 1H), 3.78−3.74 (m, 1H), 3.71–3.66 (m, 2H), 3.57 (dd, J = 15.1 Hz and 7.4 Hz, 1H), 2.04 (s, 1H, COCH3); 13C NMR (125 MHz, D2O) δ(ppm): 176.1, 71.1, 70.2, 64.2, 62.8, 57.9, 38.5, 21.5; HRMS (ESI): calcd for C8H17O4N2+ [M + H+] 205.1183, found 205.1184.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-l-mannitol dihydrochloride (A-11) (Ref. [61], reported as free base): Colourless syrup, 46 mg, 99% yield from compound A-3 (86 mg, 0.2 mmol); [α]D23 −66.7 (c 1.0 in MeOH); νmax/cm−1: 3313 (s), 2939 (s), 1115 (m), 1058 (m), 1033 (m), 1014 (m); 1H NMR (400 MHz, D2O) δ (ppm): 4.20–4.12 (m, 2H), 3.98 (dd, J = 12.5 Hz and 3.5 Hz, 1H), 3.91–3.81 (m, 2H), 3.74–3.69 (m, 1H), 3.57 (d, J = 10.5 Hz, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 76.5, 73.9, 63.4, 58.4, 58.1, 38.7; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1077.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-d-mannitol dihydrochloride (B-11) (Ref. [17]): Colourless syrup, 35 mg, 99% yield from compound B-3 (66 mg, 0.15 mmol); [α]D23 −66.7 (c 1.0 in MeOH); νmax/cm−1: 3314 (s), 2942 (s), 1113 (m), 1063 (m), 1033 (m), 1014 (m); 1H NMR (400 MHz, D2O) δ (ppm): 4.19–4.12 (m, 2H), 3.98 (dd, J = 12.6 Hz and 3.6 Hz, 1H), 3.88 (dd, J = 12.5 Hz and 3.6 Hz, 1H), 3.83 (q, J = 7.2 Hz, 1H), 3.73–3.69 (m, 1H), 3.56 (d, J = 10.5 Hz, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 76.5, 73.9, 63.4, 58.4, 58.1, 38.7; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1077.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-l-altritol dihydrochloride (C-11) (Ref. [17]): Colourless syrup, 39 mg, 99% yield from compound C-3 (73 mg, 0.17 mmol); [α]D23 −42.0 (c 1.0 in MeOH); νmax/cm−1: 3314 (s), 2923 (s), 1132 (m), 1033 (m), 1014 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.38 (t, J = 3.2 Hz, 1H), 4.34 (dd, J = 9.5 Hz and 3.7 Hz, 1H), 4.04–3.97 (m, 1H), 3.97–3.92 (m, 2H), 3.85 (ddd, J = 13.8 Hz and 7.8 Hz and 5.9 Hz, 1H), 3.62–3.53 (m, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 74.0, 69.3, 62.9, 57.3, 56.9, 39.0; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1078.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-d-altritol dihydrochloride (D-11) (Ref. [61], reported as free base): Colourless syrup, 37 mg, 99% yield from compound D-3 (69 mg, 0.2 mmol); [α]D23 +39.0 (c 0.8 in MeOH); νmax/cm−1: 3318 (s), 2923 (s), 1131 (m), 1033 (m), 983 (m); 1H NMR (500 MHz, D2O) δ(ppm): 4.38 (t, J = 2.5 Hz, 1H), 4.34 (dd, J = 9.5 Hz and 3.7 Hz, 1H), 4.06–4.01 (m, 1H), 3.98–3.93 (m, 2H), 3.86 (dd, J = 14.9 Hz and 7.9 Hz, 1H), 3.62–3.53 (m, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 74.0, 69.3, 62.9, 57.3, 56.9, 39.0; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1076.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-d-glucitol dihydrochloride (E-11a) (Ref. [17]): Colourless syrup, 38 mg, 99% yield from compound E-3a (71 mg, 0.16 mmol); [α]D23 +21.6 (c 0.5 in MeOH); νmax/cm−1: 3313 (s), 2935 (s), 1118 (m), 1063 (m), 1033 (m); 1H NMR (300 MHz, D2O) δ (ppm): 4.32–4.29 (m, 1H), 4.26 (t, J = 2.8 Hz, 1H), 4.02–3.90 (m, 3H), 3.80 (dt, J = 6.9 and 2.9 Hz, 1H), 3.54 (d, J = 6.8 Hz, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 77.5, 74.1, 64.1, 62.4, 56.9, 39.2; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1077.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-l-iditol dihydrochloride (E-11b) (Ref. [17]): Colourless syrup, 44 mg, 99% yield from compound E-3b (81 mg, 0.19 mmol); [α]D23 +5.5 (c 1.0 in MeOH); νmax/cm−1: 3322 (s), 2924 (s), 1131 (m), 1039 (m), 984 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.46 (d, J = 5.1 Hz, 1H), 4.44 (d, J = 5.1Hz, 1H), 4.22 (dt, J = 6.8 Hz and 3.5 Hz, 1H), 4.12-4.09 (m, 1H), 4.03 (dd, J = 12.2 Hz and 4.6Hz, 1H), 3.94 (dd, J = 12.2 Hz and 8.8 Hz, 1H), 3.62 (dd, J = 13.7 Hz and 6.8 Hz, 1H), 3.55–3.51(dd, J = 13.7 Hz and 6.8 Hz, 1H); 13C NMR (125 MHz, D2O) δ(ppm): 74.6, 74.2, 63.8, 58.3, 57.3, 35.9; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1076.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-l-glucitol dihydrochloride (F-11a) (Ref. [61], reported as free base): Colourless syrup, 31 mg, 99% yield from compound F-3a (57 mg, 0.13 mmol); [α]D23 −27.5 (c 0.65 in MeOH); νmax/cm−1: 3313 (s), 2935 (s), 1118 (m), 1063 (m), 1033 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.35 (s, 1H), 4.31 (s, 1H), 4.05–3.96 (m, 3H), 3.88–3.84 (m, 1H), 3.59 (d, J = 6.6 Hz, 2H); 13C NMR (125 MHz, D2O) δ(ppm): 77.4, 74.0, 64.1, 62.5, 56.8, 39.2; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1076.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-d-iditol dihydrochloride (F-11b) (Ref. [61], reported as free base): Colourless syrup, 34 mg, 99% yield from compound F-3b (62 mg, 0.14 mmol); [α]D23 −7.3 (c 0.3 in MeOH); νmax/cm−1: 3310 (s), 2936 (s), 1131 (m), 1076 (m), 1031 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.41–4.39 (m, 2H), 4.22 (dt, J = 6.7 Hz and 3.5 Hz, 1H), 4.05–3.99 (m, 2H), 3.91 (dd, J = 11.8 Hz and 8.5 Hz, 1H), 3.57 (dd, J = 13.7 Hz and 6.8 Hz, 1H), 3.47(dd, J = 13.7 Hz and 6.8 Hz, 1H); 13C NMR (125 MHz, D2O) δ (ppm): 74.7, 74.4, 63.6, 58.1, 57.4, 36.0; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1076.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-d-allitol dihydrochloride (G-11) (Ref. [61], reported as free base): Colourless syrup, 35 mg, 99% yield from compound G-3 (65 mg, 0.15 mmol); [α]D23 +8.1 (c 1.0 in MeOH); νmax/cm−1: 3322 (s), 2947 (s), 1122 (m), 1078 (m), 1034 (m), 979 (m); 1H NMR (500 MHz, D2O) δ (ppm): 4.33 (dd, J = 7.6 Hz and 5.0 Hz, 1H), 4.29 (dd, J = 4.7 Hz and 4.0 Hz, 1H), 3.96 (dd, J = 12.2 Hz and 3.5 Hz 1H), 3.92–3.84 (m, 3H) 3.61–3.53 (m, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 72.5, 70.0, 66.0, 58.6, 58.1, 38.4; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1077.
Data for 1-amino-2,5-imino-1,2,5-trideoxy-l-allitol dihydrochloride (H-11) (Ref. [61], reported as free base): Colourless syrup, 36 mg, 99% yield from compound H-3 (67 mg, 0.16 mmol); [α]D22 −7.2 (c 1.0 in MeOH); νmax/cm−1: 3313 (s), 2951 (s), 1124 (m), 1079 (m), 1034 (m), 980 (m); 1H NMR (400 MHz, D2O) δ (ppm):4.34–4.27 (m, 2H), 3.96 (dd, J = 12.0 Hz and 3.4 Hz, 1H), 3.92–3.84 (m, 3H), 3.62–3.52 (m, 2H); 13C NMR (125 MHz, D2O) δ (ppm): 72.5, 70.1, 66.1, 58.7, 58.1, 38.4; HRMS (ESI): calcd for C6H15O3N2+ [M + H+] 163.1077, found 163.1077.

4. Conclusions

In summary, a general and efficient synthetic strategy has been developed for the synthesis of 1-N-acetylamino and 1-amino pyrrolidine analogues of pochonicine (1) with l-arabino-nitrone (A), d-arabino-nitrone (B), l-lyxo-nitrone (C), d-lyxo-nitrone (D), l-xylo-nitrone (E), d-xylo-nitrone (F), l-ribo-nitrone (G) and d-ribo-nitrone (H) as the starting materials 4 and 5 steps, respectively. Glycosidase inhibition assays on a range of enzymes showed that 1-N-acetylamino pyrrolidine analogues with the same configuration as DMDP and pochonicine (1) showed powerful inhibition of β-HexNAcases and moderate inhibition of α-GalNAcase, while the other compounds showed weak or no inhibition of the tested glycosidases. This work has further examined the glycosidase inhibition of pyrrolidine analogues of pochonicine and its stereoisomers, and would be helpful for the study of potent and selective β-HexNAcase inhibitors.

Supplementary Materials

The following are available online at https://www.mdpi.com/1420-3049/25/7/1498/s1: Copies of 1H NMR, 13C NMR and 2D NMR spectra, and data of crystal structures.

Author Contributions

X.Y. performed the chemical syntheses, analysed the data and wrote the original draft; Y.S. and A.K. performed the glycosidase inhibition assay; Y.-X.L. and Y.-M.J. validated all the experimental data and revised the manuscript; A.K., G.W.J.F. and C.-Y.Y. designed the experiments and reviewed the manuscript. All authors approved the final manuscript. All authors have read and agreed to the published version of the manuscript.

Funding

Financial support from the National Natural Science Foundation of China (No. 21772206) and National Engineering Research Center for Carbohydrate Synthesis of Jiangxi Normal University is gratefully acknowledged. This work was supported in part by a Grant-in-Aid for Scientific Research (C) from the Japanese Society for the Promotion of Science (JSPS KAKENHI Grant Number JP17K08362) (AK).

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Usuki, H.; Toyo-oka, M.; Kanzaki, H.; Okuda, T.; Nitoda, T. Pochonicine, a polyhydroxylated pyrrolizidine alkaloid from fungus Pochonia suchlasporia var. suchlasporia TAMA 87 as a potent β-N-acetylglucosaminidase inhibitor. Bioorg. Med. Chem. 2009, 17, 7248–7253. [Google Scholar] [CrossRef] [PubMed]
  2. Zhu, J.-S.; Nakagawa, S.; Chen, W.; Adachi, I.; Jia, Y.-M.; Hu, X.-G.; Fleet, G.W.J.; Wilson, F.X.; Nitoda, T.; Horne, G.; et al. Synthesis of Eight Stereoisomers of Pochonicine: Nanomolar Inhibition of β-N-Acetylhexosaminidases. J. Org. Chem. 2013, 78, 10298–10309. [Google Scholar] [CrossRef] [PubMed]
  3. Stuetz, A.E.; Wrodnigg, T.M. Carbohydrate-Processing Enzymes of the Lysosome: Diseases Caused by Misfolded Mutants and Sugar Mimetics as Correcting Pharmacological Chaperones. In Advances in Carbohydrate Chemistry and Biochemistry; Baker, D.C., Ed.; Elsevier: Amsterdam, The Netherlands, 2016; Volume 73, pp. 225–302. [Google Scholar]
  4. Rast, D.M.; Baumgartner, D.; Mayer, C.; Hollenstein, G.O. Cell wall-associated enzymes in fungi. Phytochemistry 2003, 64, 339–366. [Google Scholar] [CrossRef]
  5. Merzendorfer, H.; Zimoch, L. Chitin metabolism in insects: Structure, function and regulation of chitin synthases and chitinases. J. Exp. Biol. 2003, 206, 4393–4412. [Google Scholar] [CrossRef] [Green Version]
  6. Hart, G.W.; Housley, M.P.; Slawson, C. Cycling of O-linked β-N-acetylglucosamine on nucleocytoplasmic proteins. Nature 2007, 446, 1017–1022. [Google Scholar] [CrossRef]
  7. Boyd, R.E.; Lee, G.; Rybczynski, P.; Benjamin, E.R.; Khanna, R.; Wustman, B.A.; Valenzano, K.J. Pharmacological Chaperones as Therapeutics for Lysosomal Storage Diseases. J. Med. Chem. 2013, 56, 2705–2725. [Google Scholar] [CrossRef]
  8. Vosseller, K.; Wells, L.; Lane, M.D.; Hart, G.W. Elevated nucleocytoplasmic glycosylation by O-GlcNAc results in insulin resistance associated with defects in Akt activation in 3T3-L1 adipocytes. Proc. Nat. Acad. Sci. USA 2002, 99, 5313–5318. [Google Scholar] [CrossRef] [Green Version]
  9. Yuzwa, S.A.; Macauley, M.S.; Heinonen, J.E.; Shan, X.; Dennis, R.J.; He, Y.; Whitworth, G.E.; Stubbs, K.A.; McEachern, E.J.; Davies, G.J.; et al. A potent mechanism-inspired O-GlcNAcase inhibitor that blocks phosphorylation of tau in vivo. Nat. Chem. Biol. 2008, 4, 483–490. [Google Scholar] [CrossRef]
  10. Scott, A.Y.; David, J.V. O-GlcNAc Modification and the Tauopathies: Insights from Chemical Biology. Curr. Alzheimer Res. 2009, 6, 451–454. [Google Scholar]
  11. Cecioni, S.; Vocadlo, D.J. Tools for probing and perturbing O-GlcNAc in cells and in vivo. Curr. Opin. Chem. Biol. 2013, 17, 719–728. [Google Scholar] [CrossRef]
  12. Aoyama, T.; Naganawa, H.; Suda, H.; Uotani, K.; Aoyagi, T.; Takeuchi, T. The structure of nagstatin, a new inhibitor of N-acetyl-β-d-glucosaminidase. J. Antibiot. 1992, 45, 1557–1558. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  13. Umezawa, H.; Aoyagi, T.; Komiyama, T.; Morishima, H.; Hamada, M.; Takeuchi, T. Purification and characterization of a sialidase inhibitor, siastatin, produced by streptomyces. J. Antibiot. 1974, 27, 963–969. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  14. Tatsuta, K.; Miura, S.; Ohta, S.; Gunji, H. Syntheses and glycosidase inhibiting activities of nagstatin analogs. J. Antibiot. 1995, 48, 286–288. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  15. Liang, P.H.; Cheng, W.C.; Lee, Y.L.; Yu, H.P.; Wu, Y.T.; Lin, Y.L.; Wong, C.H. Novel five-membered iminocyclitol derivatives as selective and potent glycosidase inhibitors: New structures for antivirals and osteoarthritis. Chembiochem 2006, 7, 165–173. [Google Scholar] [CrossRef]
  16. Tsou, E.-L.; Yeh, Y.-T.; Liang, P.-H.; Cheng, W.-C. A convenient approach toward the synthesis of enantiopure isomers of DMDP and ADMDP. Tetrahedron 2009, 65, 93–100. [Google Scholar] [CrossRef]
  17. Win-Mason, A.L.; Jongkees, S.A.K.; Withers, S.G.; Tyler, P.C.; Timmer, M.S.M.; Stocker, B.L. Stereoselective Total Synthesis of Aminoiminohexitols via Carbamate Annulation. J. Org. Chem. 2011, 76, 9611–9621. [Google Scholar] [CrossRef]
  18. Ayers, B.J.; Glawar, A.F.G.; Martínez, R.F.; Ngo, N.; Liu, Z.; Fleet, G.W.J.; Butters, T.D.; Nash, R.J.; Yu, C.-Y.; Wormald, M.R.; et al. Nine of 16 Stereoisomeric Polyhydroxylated Proline Amides Are Potent β-N-Acetylhexosaminidase Inhibitors. J. Org. Chem. 2014, 79, 3398–3409. [Google Scholar] [CrossRef]
  19. Glawar, A.F.G.; Martinez, R.F.; Ayers, B.J.; Hollas, M.A.; Ngo, N.; Nakagawa, S.; Kato, A.; Butters, T.D.; Fleet, G.W.J.; Jenkinson, S.F. Structural essentials for β-N-acetylhexosaminidase inhibition by amides of prolines, pipecolic and azetidine carboxylic acids. Org. Biomol. Chem. 2016, 14, 10371–10385. [Google Scholar] [CrossRef]
  20. Rountree, J.S.S.; Butters, T.D.; Wormald, M.R.; Dwek, R.A.; Asano, N.; Ikeda, K.; Evinson, E.L.; Nash, R.J.; Fleet, G.W.J. Efficient synthesis from D-lyxonolactone of 2-acetamido-1,4-imino-1,2,4-trideoxy-L-arabinitol LABNAc, a potent pyrrolidine inhibitor of hexosaminidases. Tetrahedron Lett. 2007, 48, 4287–4291. [Google Scholar] [CrossRef]
  21. Crabtree, E.V.; Martinez, R.F.; Nakagawa, S.; Adachi, I.; Butters, T.D.; Kato, A.; Fleet, G.W.J.; Glawar, A.F.G. Synthesis of the enantiomers of XYLNAc and LYXNAc: Comparison of β-N-acetylhexosaminidase inhibition by the 8 stereoisomers of 2-N-acetylamino-1,2,4-trideoxy-1,4-iminopentitols. Org. Biomol. Chem. 2014, 12, 3932–3943. [Google Scholar] [CrossRef]
  22. Tran, A.T.; Luo, B.; Jagadeesh, Y.; Auberger, N.; Désiré, J.; Nakagawa, S.; Kato, A.; Zhang, Y.; Blériot, Y.; Sollogoub, M. Synthesis of pyrrolidine-based analogues of 2-acetamidosugars as N-acetyl-d-glucosaminidase inhibitors. Carbohydr. Res. 2015, 409, 56–62. [Google Scholar] [CrossRef] [Green Version]
  23. Fleet, G.W.J.; Smith, P.W.; Nash, R.J.; Fellows, L.E.; Parekh, R.B.; Rademacher, T.W. Synthesis of 2-Acetamido-1,5-imino-1,2,5-trideoxy-d-mannitol and of 2-Acetamido-1,5-imino-1,2,5-trideoxy-d-glucitol, a Potent and Specific Inhibitor of a Number of β-N-Acetylglucosaminidases. Chem. Lett. 1986, 15, 1051–1054. [Google Scholar] [CrossRef]
  24. Glawar, A.F.G.; Best, D.; Ayers, B.J.; Miyauchi, S.; Nakagawa, S.; Aguilar-Moncayo, M.; García Fernández, J.M.; Ortiz Mellet, C.; Crabtree, E.V.; Butters, T.D.; et al. Scalable Syntheses of Both Enantiomers of DNJNAc and DGJNAc from Glucuronolactone: The Effect of N-Alkylation on Hexosaminidase Inhibition. Chem. Eur. J. 2012, 18, 9341–9359. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. de la Fuente, A.; Verdaguer, X.; Riera, A. Stereodivergent Syntheses of altro and manno Stereoisomers of 2-Acetamido-1,2-dideoxynojirimycin. Eur. J. Org. Chem. 2017, 47, 7179–7185. [Google Scholar] [CrossRef]
  26. Blériot, Y.; Tran, A.T.; Prencipe, G.; Jagadeesh, Y.; Auberger, N.; Zhu, S.; Gauthier, C.; Zhang, Y.; Désiré, J.; Adachi, I.; et al. Synthesis of 1,2-trans-2-Acetamido-2-deoxyhomoiminosugars. Org. Lett. 2014, 16, 5516–5519. [Google Scholar] [CrossRef]
  27. Li, H.; Marcelo, F.; Bello, C.; Vogel, P.; Butters, T.D.; Rauter, A.P.; Zhang, Y.; Sollogoub, M.; Blériot, Y. Design and synthesis of acetamido tri- and tetra-hydroxyazepanes: Potent and selective β-N-acetylhexosaminidase inhibitors. Bioorg. Med. Chem. 2009, 17, 5598–5604. [Google Scholar] [CrossRef]
  28. Marcelo, F.; He, Y.; Yuzwa, S.A.; Nieto, L.; Jiménez-Barbero, J.; Sollogoub, M.; Vocadlo, D.J.; Davies, G.D.; Blériot, Y. Molecular Basis for Inhibition of GH84 Glycoside Hydrolases by Substituted Azepanes: Conformational Flexibility Enables Probing of Substrate Distortion. J. Am. Chem. Soc. 2009, 131, 5390–5392. [Google Scholar] [CrossRef] [Green Version]
  29. Mondon, M.; Hur, S.; Vadlamani, G.; Rodrigues, P.; Tsybina, P.; Oliver, A.; Mark, B.L.; Vocadlo, D.J.; Blériot, Y. Selective trihydroxyazepane NagZ inhibitors increase sensitivity of Pseudomonas aeruginosa to β-lactams. Chem. Commun. 2013, 49, 10983–10985. [Google Scholar] [CrossRef]
  30. Glawar, A.F.G.; Jenkinson, S.F.; Thompson, A.L.; Nakagawa, S.; Kato, A.; Butters, T.D.; Fleet, G.W.J. 3-Hydroxyazetidine Carboxylic Acids: Non-Proteinogenic Amino Acids for Medicinal Chemists. Chemmedchem 2013, 8, 658–666. [Google Scholar] [CrossRef]
  31. Liu, Z.; Jenkinson, S.F.; Kato, A.; Nakagawa, S.; Wormald, M.R.; Yu, C.-Y.; Fleet, G.W.J. 3-Azidoazetidines as the first scaffolds for β-amino azetidine carboxylic acid peptidomimetics: Azetidine iminosugars containing an acetamido group do not inhibit β-N-acetylhexosaminidases. Tetrahedron Asymmetry 2016, 27, 872–881. [Google Scholar] [CrossRef]
  32. Harit, V.K.; Ramesh, N.G. Amino-functionalized iminocyclitols: Synthetic glycomimetics of medicinal interest. Rsc Advances 2016, 6, 109528–109607. [Google Scholar] [CrossRef]
  33. Kitamura, Y.; Koshino, H.; Nakamura, T.; Tsuchida, A.; Nitoda, T.; Kanzaki, H.; Matsuoka, K.; Takahashi, S. Total synthesis of the proposed structure for pochonicine and determination of its absolute configuration. Tetrahedron Lett. 2013, 54, 1456–1459. [Google Scholar] [CrossRef]
  34. Salunke, R.V.; Ramesh, N.G. A Concise Total Synthesis of the Stereoisomers of (–)-Pochonicine. Eur. J. Org. Chem. 2016, 654–657. [Google Scholar] [CrossRef]
  35. Martinez, S.T.; Belouezzane, C.; Pinto, A.C.; Glasnov, T. Synthetic Strategies towards the Azabicyclo 3.3.0 -Octane Core of Natural Pyrrolizidine Alkaloids. An Overview. Org. Prep. Proced. Int. 2016, 48, 223–253. [Google Scholar] [CrossRef]
  36. Robertson, J.; Stevens, K. Pyrrolizidine alkaloids: Occurrence, biology, and chemical synthesis. Nat. Prod. Rep. 2017, 34, 62–89. [Google Scholar] [CrossRef] [PubMed]
  37. O’Keefe, S.; Roebuck, Q.P.; Nakagome, I.; Hirono, S.; Kato, A.; Nash, R.; High, S. Characterizing the selectivity of ER α-glucosidase inhibitors. Glycobiology 2019, 29, 530–542. [Google Scholar] [CrossRef] [Green Version]
  38. Ferhati, X.; Matassini, C.; Fabbrini, M.G.; Goti, A.; Morrone, A.; Cardona, F.; Moreno-Vargas, A.J.; Paoli, P. Dual targeting of PTP1B and glucosidases with new bifunctional iminosugar inhibitors to address type 2 diabetes. Bioorg. Chem. 2019, 87, 534–549. [Google Scholar] [CrossRef]
  39. Yang, L.-F.; Shimadate, Y.; Kato, A.; Li, Y.-X.; Jia, Y.-M.; Fleet, G.W.J.; Yu, C.-Y. Synthesis and glycosidase inhibition of N-substituted derivatives of DIM. Org. Biomol. Chem. 2020, 18, 999–1011. [Google Scholar] [CrossRef]
  40. Suzuki, K.; Nakahara, T.; Kanie, O. 3,4-dihydroxypyrrolidine as glycosidase inhibitor. Curr. Top. Med. Chem. 2009, 9, 34–57. [Google Scholar] [CrossRef]
  41. Takaoka, Y.; Kajimoto, T.; Wong, C.H. Inhibition of N-acetylglucosaminyl transfer enzymes: Chemical-enzymic synthesis of new five-membered acetamido azasugars. J. Org. Chem. 1993, 58, 4809–4812. [Google Scholar] [CrossRef]
  42. Liu, J.J.; Numa, M.M.D.; Liu, H.T.; Huang, S.J.; Sears, P.; Shikhman, A.R.; Wong, C.H. Synthesis and High-Throughput Screening of N-Acetyl-β-hexosaminidase Inhibitor Libraries Targeting Osteoarthritis. J. Org. Chem. 2004, 69, 6273–6283. [Google Scholar] [CrossRef] [PubMed]
  43. Li, Y.-X.; Kinami, K.; Hirokami, Y.; Kato, A.; Su, J.-K.; Jia, Y.-M.; Fleet, G.W.J.; Yu, C.-Y. Gem-difluoromethylated and trifluoromethylated derivatives of DMDP-related iminosugars: Synthesis and glycosidase inhibition. Org. Biomol. Chem. 2016, 14, 2249–2263. [Google Scholar] [CrossRef] [PubMed]
  44. Song, Y.-Y.; Kinami, K.; Kato, A.; Jia, Y.-M.; Li, Y.-X.; Fleet, G.W.J.; Yu, C.-Y. First total synthesis of (+)-broussonetine W: Glycosidase inhibition of natural product & analogs. Org. Biomol. Chem. 2016, 14, 5157–5174. [Google Scholar] [PubMed]
  45. Cheng, B.; Hirokami, Y.; Li, Y.-X.; Kato, A.; Jia, Y.-M.; Yu, C.-Y. Synthesis and glycosidase inhibition of C-7 modified casuarine derivatives. Chin. Chem. Lett. 2017, 28, 1701–1704. [Google Scholar] [CrossRef]
  46. Wu, Q.-K.; Kinami, K.; Kato, A.; Li, Y.-X.; Jia, Y.-M.; Fleet, G.W.J.; Yu, C.-Y. Synthesis and Glycosidase Inhibition of Broussonetine M and Its Analogues. Molecules 2019, 24, 3712. [Google Scholar] [CrossRef] [Green Version]
  47. Revuelta, J.; Cicchi, S.; Goti, A.; Brandi, A. Enantiopure Cyclic Nitrones: A Useful Class of Building Blocks for Asymmetric Syntheses. Synthesis 2007, 2007, 485–504. [Google Scholar] [CrossRef]
  48. Murahashi, S.-I.; Imada, Y. Synthesis and Transformations of Nitrones for Organic Synthesis. Chem. Rev. 2019, 119, 4684–4716. [Google Scholar] [CrossRef]
  49. Holzapfel, C.W.; Crous, R. Synthesis and reactions of chiral cyclic nitrones derived from d-ribose. Heterocycles 1998, 48, 1337–1342. [Google Scholar] [CrossRef]
  50. Pillard, C.; Desvergnes, V.; Py, S. Diastereoselective addition of alkynylalanes to carbohydrate-derived nitrones. Tetrahedron Lett. 2007, 48, 6209–6213. [Google Scholar] [CrossRef]
  51. Wang, W.-B.; Huang, M.-H.; Li, Y.-X.; Rui, P.-X.; Hu, X.-G.; Zhang, W.; Su, J.-K.; Zhang, Z.-L.; Zhu, J.-S.; Xu, W.-H.; et al. A Practical Synthesis of Sugar-derived Cyclic Nitrones: Powerful Synthons for the Synthesis of Iminosugars. Synlett 2010, 3, 488–492. [Google Scholar]
  52. Messire, G.; Massicot, F.; Vallee, A.; Vasse, J.-L.; Behr, J.-B. Aza-Henry Reaction with Nitrones, an Under-Explored Transformation. Eur. J. Org. Chem. 2019, 7, 1659–1668. [Google Scholar] [CrossRef]
  53. Cheng, W.-C.; Wang, J.-H.; Yun, W.-Y.; Li, H.-Y.; Hu, J.-M. Rapid preparation of (3R,4S,5R) polyhydroxylated pyrrolidine-based libraries to discover a pharmacological chaperone for treatment of Fabry disease. Eur. J. Med. Chem. 2017, 126, 1–6. [Google Scholar] [CrossRef] [PubMed]
  54. Hu, X.-G.; Bartholomew, B.; Nash, R.J.; Wilson, F.X.; Fleet, G.W.J.; Nakagawa, S.; Kato, A.; Jia, Y.-M.; Well, R.V.; Yu, C.-Y. Synthesis and Glycosidase Inhibition of the Enantiomer of (-)-Steviamine, the First Example of a New Class of Indolizidine Alkaloid. Org. Lett. 2010, 12, 2562–2565. [Google Scholar] [CrossRef] [PubMed]
  55. Li, Y.-X.; Huang, M.-H.; Yamashita, Y.; Kato, A.; Jia, Y.-M.; Wang, W.-B.; Fleet, G.W.J.; Nash, R.J.; Yu, C.-Y. L-DMDP, L-homoDMDP and their C-3 fluorinated derivatives: Synthesis and glycosidase-inhibition. Org. Biomol. Chem. 2011, 9, 3405–3414. [Google Scholar] [CrossRef]
  56. Tice, C.M.; Ganem, B. The chemistry of naturally occurring polyamines. 6. Efficient syntheses of N1- and N8-acetylspermidine. J. Org. Chem. 1983, 48, 2106–2108. [Google Scholar] [CrossRef]
  57. Welter, A.; Jadot, J.; Dardenne, G.; Marlier, M.; Casimir, J. 2,5-Dihydroxymethyl 3,4-dihydroxypyrrolidine dans les feuilles de Derris elliptica. Phytochemistry 1976, 15, 747–749. [Google Scholar] [CrossRef]
  58. Kessler, M.; Acuto, O.; Storelli, C.; Murer, H.; Semenza, G.A. A modified procedure for the rapid preparation of efficiently transporting vesicles from small intestinal brush border membranes. Their use in investigating some properties of d-glucose and choline transport systems. Biochim. Biophys. Acta 1978, 506, 136–154. [Google Scholar] [CrossRef]
  59. Ayers, B.J.; Ngo, N.; Jenkinson, S.F.; Martínez, R.F.; Shimada, Y.; Adachi, I.; Weymouth-Wilson, A.C.; Kato, A.; Fleet, G.W.J. Glycosidase Inhibition by All 10 Stereoisomeric 2,5-Dideoxy-2,5-iminohexitols Prepared from the Enantiomers of Glucuronolactone. J. Org. Chem. 2012, 77, 7777–7792. [Google Scholar] [CrossRef]
  60. Cheng, T.-J.R.; Chan, T.-H.; Tsou, E.-L.; Chang, S.-Y.; Yun, W.-Y.; Yang, P.-J.; Wu, Y.-T.; Cheng, W.-C. From Natural Product-Inspired Pyrrolidine Scaffolds to the Development of New Human Golgi α-Mannosidase II Inhibitors. Chem. Asian J. 2013, 8, 2600–2604. [Google Scholar] [CrossRef]
  61. Cheng, W.-C.; Wang, J.-H.; Li, H.-Y.; Lu, S.-J.; Hu, J.-M.; Yun, W.-Y.; Chiu, C.-H.; Yang, W.-B.; Chien, Y.-H.; Hwu, W.-L. Bioevaluation of sixteen ADMDP stereoisomers toward alpha-galactosidase A: Development of a new pharmacological chaperone for the treatment of Fabry disease and potential enhancement of enzyme replacement therapy efficiency. Eur. J. Med. Chem. 2016, 123, 14–20. [Google Scholar] [CrossRef]
  62. Takebayashi, M.; Hiranuma, S.; Kanie, Y.; Kajimoto, T.; Kanie, O.; Wong, C.H. A Versatile Synthetic Strategy for the Preparation and Discovery of New Iminocyclitols as Inhibitors of Glycosidases. J. Org. Chem. 1999, 64, 5280–5291. [Google Scholar] [CrossRef]
  63. Kang, S.H.; Ryu, D.H. Intramolecular cyclization of C2 symmetric and meso-iodo amino alcohols: A synthetic approach to azasugars. Tetrahedron Lett. 1997, 38, 607–610. [Google Scholar] [CrossRef]
  64. Ganesan, M.; Madhukarrao, R.V.; Ramesh, N.G. Design and synthesis of new amino-modified iminocyclitols: Selective inhibitors of α-galactosidase. Org. Biomol. Chem. 2010, 8, 1527–1530. [Google Scholar] [CrossRef] [PubMed]
Sample Availability: Samples of the compounds are not available from the authors.
Molecules 25 01498 g001 550
Figure 1. Naturally occurring potent N-acetylhexosaminidases (HexNAcase) inhibitors.
Figure 1. Naturally occurring potent N-acetylhexosaminidases (HexNAcase) inhibitors.
Molecules 25 01498 g001
Molecules 25 01498 g002 550
Figure 2. Examples of synthetic HexNAcase inhibitors.
Figure 2. Examples of synthetic HexNAcase inhibitors.
Molecules 25 01498 g002
Molecules 25 01498 g003 550
Figure 3. Polyhydroxylated nitrones used in building pyrrolidine cores.
Figure 3. Polyhydroxylated nitrones used in building pyrrolidine cores.
Molecules 25 01498 g003
Molecules 25 01498 sch001 550
Scheme 1. Synthesis of compound A-11 from l-arabino-nitrone (A).
Scheme 1. Synthesis of compound A-11 from l-arabino-nitrone (A).
Molecules 25 01498 sch001
Molecules 25 01498 sch002 550
Scheme 2. Synthesis of monoacetylated compound A-10.
Scheme 2. Synthesis of monoacetylated compound A-10.
Molecules 25 01498 sch002
Table
Table 1. 1-Amino and 1-N-acetylamino pyrrolidine analogues synthesized from cyclic nitrones.
Table 1. 1-Amino and 1-N-acetylamino pyrrolidine analogues synthesized from cyclic nitrones.
EntryCyclic NitroneHydroxylamineDiamineMonoacetylated Pyrrolidine1-Amino Product1-N-Acetylamino Product
1 Molecules 25 01498 i001 Molecules 25 01498 i002 Molecules 25 01498 i003 Molecules 25 01498 i004 Molecules 25 01498 i005 Molecules 25 01498 i006
2 Molecules 25 01498 i007 Molecules 25 01498 i008 Molecules 25 01498 i009 Molecules 25 01498 i010 Molecules 25 01498 i011 Molecules 25 01498 i012
3 Molecules 25 01498 i013 Molecules 25 01498 i014 Molecules 25 01498 i015 Molecules 25 01498 i016 Molecules 25 01498 i017 Molecules 25 01498 i018
4 Molecules 25 01498 i019 Molecules 25 01498 i020 Molecules 25 01498 i021 Molecules 25 01498 i022 Molecules 25 01498 i023 Molecules 25 01498 i024
5 Molecules 25 01498 i025 Molecules 25 01498 i026 Molecules 25 01498 i027 Molecules 25 01498 i028 Molecules 25 01498 i029 Molecules 25 01498 i030
Molecules 25 01498 i031 Molecules 25 01498 i032 Molecules 25 01498 i033 Molecules 25 01498 i034
6 Molecules 25 01498 i035 Molecules 25 01498 i036 Molecules 25 01498 i037 Molecules 25 01498 i038 Molecules 25 01498 i039 Molecules 25 01498 i040
Molecules 25 01498 i041 Molecules 25 01498 i042 Molecules 25 01498 i043 Molecules 25 01498 i044
7 Molecules 25 01498 i045 Molecules 25 01498 i046 Molecules 25 01498 i047 Molecules 25 01498 i048 Molecules 25 01498 i049 Molecules 25 01498 i050
8 Molecules 25 01498 i051 Molecules 25 01498 i052 Molecules 25 01498 i053 Molecules 25 01498 i054 Molecules 25 01498 i055 Molecules 25 01498 i056
a Yield of the corresponding hydroxylamines starting from cyclic nitrones; b Total yield in 2 steps starting from hydroxylamines to diamines; c Yield of monoacetylated pyrrolidines from the corresponding diamines; d Hydrogenation yield of 1-amino products from diamines; e Hydrogenation yield of 1-N-acetylamino products from the corresponding monoacetylated pyrrolidines.
Table
Table 2. Concentrations of 1-N-acetylamino pyrrolidine analogues giving 50% inhibition of various enzymes.
Table 2. Concentrations of 1-N-acetylamino pyrrolidine analogues giving 50% inhibition of various enzymes.
EnzymeIC50 (μM)
Molecules 25 01498 i057 Molecules 25 01498 i058 Molecules 25 01498 i059 Molecules 25 01498 i060 Molecules 25 01498 i061 Molecules 25 01498 i062 Molecules 25 01498 i063 Molecules 25 01498 i064 Molecules 25 01498 i065 Molecules 25 01498 i066 Molecules 25 01498 i067
A-104·HClC-10D-10E-10aF-10aE-10bF-10bG-10H-10Pochonicine (1)
α-Glucosidase
 YeastNIa (0.5%)b450NI (0%)NI (0%)NI (0%)NI (6.91%)NI (0%)NI (0%)NI (0%)NI (0%)—c
 Rice5681000NI (18.2%)NI (26.1%)NI (15.9%)NI (12.5%)NI (0%)NI (0%)NI (0%)NI (5.6%)—
 Rat intestinal maltase92211NI (35.4%)NI (9.24%)NI (12.7%)NI (14.6%)NI (0%)NI (12.7%)NI (16.0%)NI (13.6%)—
β-Glucosidase
 AlmondNI (16.1%)170NI (0%)NI (41.0%)NI (37.9%)NI (11.6%)NI (1.2%)NI (19.8%)NI (15.3%)NI (17.1%)—
 Bovine liverNI (2.0%)NI (33.8%)NI (1.0%)NI (2.4%)NI (4.1%)NI (0.3%)NI (0%)NI (0%)NI (0%)NI (1.7%)—
α-Galactosidase
 Coffee beansNI (0%)NI (1.1%)NI (0.36%)380NI (0.36%)NI (0.36%)NI (0%)NI (0%)NI (0%)NI (0%)—
β-Galactosidase
 Bovine liverNI (1.6%)511NI (1.3%)NI (3.3%)NI (4.9%)NI (4.2%)NI (3.9%)NI (4.2%)NI (2.9%)NI (3.6%)—
α-Mannosidase
 Jack beanNI (0%)NI (0%)NI (0.4%)205NI (0.7%)NI (0%)NI (3.8%)NI (1.6%)NI (4.0%)NI (9.5%)—
β-Mannosidase
 SnailNI (0%)296NI (0%)NI (0%)NI (25.8%)NI (3.23%)NI (0%)NI (0%)NI (0%)NI (0%)—
α-l-Fucosidase
 Bovine kidneyNI (0%)NI (1.5%)NI (4.9%)NI (1.7%)NI (41.7%)NI (8.4%)NI (20.5%)NI (3.9%)NI (11.5%)NI (22.3%)—
α,α-Trehalase
 Porcine kidneyNI (4.3%)NI (20.3%)NI (0%)NI (0%)NI (5.6%)NI (3.0%)NI (0%)NI (0%)NI (5.6%)NI (40.9%)—
Amyloglucosidase
 A. nigerNI (5.5%)NI (40.6%)NI (1.9%)NI (3.8%)NI (4.0%)NI (2.8%)NI (6.4%)NI (5.7%)NI (7.6%)NI (8.3%)—
α-l-Rhamnosidase
 P. decumbensNI (10.1%)NI (6.5%)NI (3.6%)NI (8.3%)NI (11.8%)NI (5.3%)NI (30.8%)NI (3.0%)NI (9.5%)NI (7.7%)—
β-Glucuronidase
 E. coliNI (0%)NI (0.6%)NI (0.3%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)—
 Bovine liverNI (4.6%)NI (1.7%)NI (0%)NI (1.3%)NI (0%)NI (0%)NI (4.3%)NI (0%)NI (0%)NI (0%)—
β-N Acetylhexosaminidase
 Bovine liver9434.7NI (36.3%)2.895NI (23.2%)NI (38.4%)652299NI (33.7%)0.021 [2]
 HL60NI (12.7%)34NI (3.9%)10591NI (0%)NI (4.5%)NI (16.6%)NI (18.8%)NI (0.3%)0.018 [2]
 Jack bean2020.211290.1210NI (42%)11598262410.0016 [2]
 A. oryzae——————————0.33 [2]
 Human placenta——————————0.012 [2]
β-N-Acetylgalactosaminidase
 HL60NI (13.0%)9.5NI (1.4%)8.8490NI (0.9%)NI (10.1%)NI (36.1%)NI (15.9%)NI (5.3%)0.049 [2]
α-N-Acetylgalactosaminidase
 Chicken liverNI (0%)NI (3.3%)NI (6.1%)65.3NI (1.4%)NI (0%)NI (0.5%)NI (2.3%)NI (2.3%)NI (6.1%)NI (9.0%) [2]
a NI: No Inhibition (less than 50% at 1000 μM); b ( ): Inhibition % at 1000 μM; c —: Not determined.
Table
Table 3. Concentrations of 1-amino pyrrolidine analogues giving 50% inhibition of various enzymes.
Table 3. Concentrations of 1-amino pyrrolidine analogues giving 50% inhibition of various enzymes.
EnzymeIC50 (μM)
Molecules 25 01498 i068 Molecules 25 01498 i069 Molecules 25 01498 i070 Molecules 25 01498 i071 Molecules 25 01498 i072 Molecules 25 01498 i073 Molecules 25 01498 i074 Molecules 25 01498 i075 Molecules 25 01498 i076 Molecules 25 01498 i077
A-11B-11C-11D-11E-11F-11aE-11bF-11bG-11H-11
α-Glucosidase
 YeastNIa (0%)bNI (42.7%)NI (9.55%)NI (4.52%)NI (9.55%)NI (10.1%)NI (0%)NI (0%)NI (0%)NI (0%)
 Rice286725NI (0%)NI (8.4%)NI (15.7%)NI (31.5%)NI (12.9%)NI (16.9%)NI (11.8%)NI (3.4%)
 Rat intestinal maltase68251NI (37.1%)NI (4.02%)NI (9.95%)NI (48.0%)NI (10.9%)NI (20.8%)NI (%2.64)NI (10.1%)
β-Glucosidase
 AlmondNI (0%)NI (44.4%)NI (13.0%)419NI (12.4%)NI (6.0%)NI (4.8%)NI (13.5%)NI (2.3%)NI (18.8%)
 Bovine liverNI (0%)NI (16.2%)NI (0.7%)NI (3.1%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (3.8%)
α-Galactosidase
 Coffee beansNI (0%)NI (0%)NI (1.9%)NI (42.2%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)
β-Galactosidase
 Bovine liverNI (1.0%)NI (36.3%)NI (2.6%)NI (6.5%)NI (2.0%)NI (0%)NI (0%)NI (2.6%)NI (0%)NI (4.2%)
α-Mannosidase
 Jack beanNI (0%)NI (0%)NI (0%)54NI (0.32%)NI (0.97%)NI (12.1%)NI (0%)NI (5.19%)NI (3.81%)
β-Mannosidase
 SnailNI (0%)NI (0%)NI (3.35%)NI (2.23%)NI (2.6%)NI (1.9%)NI (0%)NI (0%)NI (0%)NI (1.2%)
α-l-Fucosidase
 Bovine kidneyNI (13.1%)NI (0%)NI (39.9%)NI (8.0%)NI (21.2%)NI (46.4%)NI (19.9%)NI (16.2%)NI (16.2%)NI (16.2%)
α,α-Trehalase
 Porcine kidneyNI (0%)NI (4.7%)NI (0.9%)NI (0%)NI (4.7%)NI (0%)NI (2.2%)NI (0.6%)NI (3.8%)NI (0%)
Amyloglucosidase
 A. nigerNI (0.9%)589NI (0%)NI (0.7%)NI (9.1%)NI (0%)NI (0%)NI (0%)NI (1.1%)NI (2.1%)
α-l-Rhamnosidase
 P. decumbensNI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (1.6%)NI (1.6%)NI (1.3%)
β-Glucuronidase
 E. coliNI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)
 Bovine liverNI (0.8%)NI (0%)NI (0%)NI (0%)NI (0%)NI (5.7%)NI (4.1%)NI (0%)NI (0%)NI (4.9%)
β-N-Acetylhexosaminidase
 Bovine liverNI (2.1%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (0%)NI (1.6%)NI (1.6%)NI (1.3%)
 HL60NI (0%)NI (0%)NI (0%)NI (3.1%)NI (1.3%)NI (2.7%)NI (0%)NI (0%)NI (0%)NI (0%)
 Jack beanNI (21.1%)NI (14.5%)NI (14.5%)99NI (19.3%)NI (30.7%)NI (23.5%)264NI (29.5%)NI (7.2%)
β-N-Acetylgalactosaminidase
 HL60NI (5.2%)NI (3.8%)NI (2.9%)NI (14.8%)NI (4.8%)NI (6.0%)NI (11.9%)NI (8.1%)NI (1.9%)NI (0%)
α-N-Acetylgalactosaminidase
 Chicken liverNI (4.7%)NI (7.5%)NI (0.5%)44NI (0%)NI (5.1%)NI (0%)NI (0%)NI (13.6%)NI (2.8%)
a NI: No Inhibition (less than 50% at 1000 μM); b ( ): Inhibition % at 1000 μM.

Share and Cite

MDPI and ACS Style

Yan, X.; Shimadate, Y.; Kato, A.; Li, Y.-X.; Jia, Y.-M.; Fleet, G.W.J.; Yu, C.-Y. Synthesis of Pyrrolidine Monocyclic Analogues of Pochonicine and Its Stereoisomers: Pursuit of Simplified Structures and Potent β-N-Acetylhexosaminidase Inhibition. Molecules 2020, 25, 1498. https://doi.org/10.3390/molecules25071498

AMA Style

Yan X, Shimadate Y, Kato A, Li Y-X, Jia Y-M, Fleet GWJ, Yu C-Y. Synthesis of Pyrrolidine Monocyclic Analogues of Pochonicine and Its Stereoisomers: Pursuit of Simplified Structures and Potent β-N-Acetylhexosaminidase Inhibition. Molecules. 2020; 25(7):1498. https://doi.org/10.3390/molecules25071498

Chicago/Turabian Style

Yan, Xin, Yuna Shimadate, Atsushi Kato, Yi-Xian Li, Yue-Mei Jia, George W. J. Fleet, and Chu-Yi Yu. 2020. "Synthesis of Pyrrolidine Monocyclic Analogues of Pochonicine and Its Stereoisomers: Pursuit of Simplified Structures and Potent β-N-Acetylhexosaminidase Inhibition" Molecules 25, no. 7: 1498. https://doi.org/10.3390/molecules25071498

Article Metrics

Back to TopTop