Research Article | Open Access
Thio-β-D-glucosides: Synthesis and Evaluation as Glycosidase Inhibitors and Activators
Structurally simple 1-thio-β-D-glucopyranosides were synthesized and tested as potential inhibitors toward several fungal glycosidases from Aspergillus oryzae and Penicillium canescens. Significant selective inhibition was observed for α- and β-glucosidases, while a weak to moderate activation for α- and β-galactosidases.
Thioglycosides are the hydrolysis- and metabolism-resistant synthetic S-analogs of natural O-glycosides. They attracted recently a rapidly increasing attention as competitive inhibitors of glycosidases and other enzymes involved in a variety of biochemical processes [1, 2] related in particular to metabolic disorders and diseases, such as diabetes [1, 3], to inflammations [2, 4] and viral or bacterial infections [5–17], including tuberculosis [9, 10], and to cancer [2, 18–20]. However, surprisingly little is known about the inhibition of glucosidases by 1-thio-β-D-glucosides so far [21–24] (see the discussion). In a search for simple, readily accessible, and efficient glycosidase inhibitors [25–27], we designed, prepared, and assayed a series of structurally simple 1-thio-β-D-glucosides.
2. Results and Discussion
2.1. Synthesis of 1-Thio-β-D-glucopyranosides
A series of aryl and alkyl 1-thio-β-D-glucopyranosides 2a–2i was synthesized according to Scheme 1. In addition to the model compounds 2a–2f with simple aryl and alkyl aglycones, we included the phenanthroline derivative 2g, which was found to possess activity towards some glycosidases . Compounds 2h–2i were designed as disaccharide analogues in view of the significant inhibitory activity of the 1,2-cyclohexanedicarboxylic acid derivatives towards fungal glycosidases [25, 26].
The structures of all products and intermediates were determined from the data of 1H NMR and 13C NMR, including COSY and HMQC techniques. Thanks to a bias of the conformational equilibria towards the all-equatorial form of the pyranose ring, the configurational assignment was rather straightforward: large spin-spin coupling constants (9–11 Hz) indicated a trans-diaxial orientation of the corresponding vicinal protons, while all other relative positions (axial-equatorial and equatorial-equatorial) resulted in small couplings between them (2–4 Hz).
2.2. Glycosidase Inhibitory Activity
The synthesized compounds have been assayed for enzyme inhibitory activity against several glycosidases in multienzyme complexes isolated from fungi Penicillium canescens and Aspergillus oryzae [28–32]. The multienzyme complex from P. canescens contains α-D- and β-D-galactosidase, β-D-glucosidase, and α-L- and β-D-fucosidase [28–31]. The multienzyme complex from A. oryzae contains α-D- and β-D-galactosidase, α-D- and β-D-glucosidase, and α-L- and β-D-fucosidase . All assays have been performed in a standard way  by monitoring spectrophotometrically at 400 nm the release of p-nitrophenol from the corresponding p-nitrophenyl glycosides. The inhibitory activity of studied compounds was found to depend on the structure of inhibitor, the type of enzyme, and the particular source of enzyme.
Usually, the structural analogues of carbohydrates with certain configuration are expected to influence activity of the corresponding enzymes (“like inhibits like”). Thus, 4-nitrophenyl 1-thio-β-D-glucoside was used to study binding to β-D-glucosidase from barley and rice and was shown to be a competitive inhibitor . As another example, a series of alkyl/aryl 1-thio-β-D-galactosides was designed and studied as inhibitors of β-D-galactosidase from E. coli . Similarly, we tried the 1-thio-β-D-glucosides 2a–2i first of all as possible inhibitors against β-D-glucosidases. In the preliminary tests, all studied compounds demonstrated indeed a substantial inhibitory activity toward fungal β-D-glucosidases from both P. canescens and A. oryzae strains (Figures 1 and 2). It is worth mentioning that compounds 2a–2f were 10-fold more potent in the case of β-D-glucosidase from A. oryzae than methyldeoxynojirimycin at 85% inhibition .
It was found previously that the inhibitors of β-D-glucosidase may also possess the α-D-glucosidase inhibitory activity [25, 26, 35]. For example, N-substituted aminomethyl-β-D-glucopyranosides inhibited yeast and rat intestinal α-D-glucosidases . In this study, compounds 2a–2h showed only weak inhibition of α-D-glucosidase from A. oryzae (Figure 1).
Quite unusually, 1-thio-β-D-glucosides revealed a weak to moderate ability to activate α- and β-D-galactosidases (Figures 1 and 2), especially α-D-galactosidase from P. canescens (Figure 2). The strongest activation was observed in the case of phenanthrolyl-1-thio-β-D-glucoside 2g (Figures 1 and 2). Some derivatives of 1,10-phenanthroline were found earlier to activate β-D-galactosidases, while other derivatives acted as inhibitors . Towards the fungal β-D-glucosidases, all 1,10-phenanthrolines were activators, with exception of thioglucoside 2g, which was an inhibitor . Noteworthy, the nitrogen analog of 2g, N-(1,10-phenanthrolin-5-yl)-β-D-glucopyranosylamine was recently synthesized, but no data on enzyme inhibition or activation were obtained yet . The mechanism of activation is currently unknown. It may be a result of an allosteric effect in these enzymes  or of a change in their hydration . The activation of α/β-D-galactosidases may be interesting for treatment of certain diseases and merits further studies [37, 38].
The analysis of the Lineweaver-Burk plots indicated that compounds 2a–2i inhibited the β-D-glucosidases competitively. Their inhibition parameters are presented in Table 1. The values of are systematically larger for the enzyme from A. oryzae as compared to P. canescens (by factors 32–35 for 2a, 2b, 2d, and 2g; and by factors 17–20 for 2c, 2e, 2f, and 2h). Similar ratios were obtained earlier for the inhibition of these enzymes by derivatives of cyclohexane-1,2-dicarboxylic acids . This is apparently due to structural differences of the enzymes, which result also in different values of Michaelis constant: μM (A. oryzae) and μM (P. canescens) . Interestingly, higher values of were found also for β-D-galactosidase from A. oryzae as compared to enzyme from P. canescens .
| = 1530 μM (for -D-glucosidase from fungus Stachybotrys atra) .|
The previously available data on 1-thio-β-D-glucopyranosides as glycosidase inhibitors were limited to the derivatives with the following aglycones: phenyl (2a), μM (Stachybotrys atra) ; 4-nitrophenyl, μM (rice) and μM (barley) ; benzo-1,4-quinone, μM (A. faecalis); chlorobenzo-1,4-quinone, μM (A. faecalis) ; and 2-(3-methyl-3H-diazirine-3-yl)-ethyl, μM (sweet almond) . Although thioglycosides are usually resistant to hydrolysis, it was found also that with some specific aglycones they may be hydrolysable and can serve as glycosidase substrates . It is interesting to compare our results (Table 1) to the data for some known inhibitors of β-D-glucosidases: 1-deoxynojirimycin, μM (Aspergillus wentii); μM (Agrobacterium faecalis); N-methyl-1-deoxynojirimycin, μM (sweet almond); and N-hexyl-1-deoxynojirimycin, μM (calf liver cytosolic) and μM (calf spleen lysosomal) . From this comparison, compounds 2a–2h are moderately strong β-D-glucosidase inhibitors.
The comparison of thioglucosides 2a–2i allows a preliminary elucidation of the effect of aglycon upon their inhibitory activity. It is known that the presence of alkyl groups and other lipophilic moieties often increases the efficiency of glycosidase inhibitors. This effect, mostly towards β-glucosidases, was observed for derivatives of 1-deoxynojirimycin and other iminosugars, alkyl glycosides, carbasugars, and some other inhibitors. It was explained by additional stabilization of the alkyl chains or aromatic residues by the hydrophobic pocket or cleft within or in vicinity of the enzyme binding site (for the detailed discussion see [25, 42, 43] and references therein; in addition see [37, 44, 45]). The results of our study (Table 1) seem to confirm the general regularities observed earlier. Thus, the most potent inhibitors of β-D-glucosidases, compounds 2f and 2g, have large lipophilic aglycones, while the thioglucoside 2i with the most polar aglycone in this series shows almost negligible inhibition. However, the dependence of inhibition on the aglycone’s lipophilicity is not straightforward: despite their large aromatic aglycones, the thioglucosides 2a–2e produced much weaker inhibition than 2f and 2 g (see also discussion in ). The influence of the aglycone shape remains also uncertain. The best results were achieved both for the narrow alkyl chain (2f) and for the wide polyaromatic moiety (2g).
3. Experimental Section
The chemicals used in this study were purchased from commercial sources (Sigma-Aldrich, TCI, and Across Organics) and used without additional purification. All solvents were purified by conventional techniques prior to use. Column chromatography was performed on silica gel (40–75 μm, Sorbent Technologies) and aluminum oxide (activated basic, 58Å, Aldrich). The reactions were monitored by TLC on silica gel plates, Analtech Inc (visualization by staining with sulfuric acid followed by heating or with ).
1H NMR and 13C NMR spectra were acquired on JEOL ECA-600 NMR-spectrometer (600 MHz for 1H and 150 MHz for 13C) with spinning at rt. 1H-1H-COSY and 1H-13C-HMQC techniques were used to assign the signals. High resolution mass spectra (HRMS) were obtained on a JEOL AccuTOF time-of-flight mass spectrometer (Peabody, MA) coupled with an Ionsense DART open-air ionization source (Saugus, MA). The instrument was tuned to a resolving power of 7,000 with reserpine directly infused into the electrospray ionization source; this provided a stable ion current to tune the time-of-flight parameters. Samples were introduced into the DART sample gap with a glass melting point capillary by first dipping the closed end of the capillary into the sample then immediately placing it into the helium metastable beam. The helium gas temperature was set to 250°C to aid in the desorption of the analyte from the capillary. The samples were held in the sample gap for 10–15 seconds to acquire several mass spectra to average for an accurate m/z assignment. Optical rotation was measured for solutions in a 5 cm cell with an automatic polarimeter Autopol I. A Beckman Du-65 spectrometer was used for the enzymatic hydrolysis studies.
3.1. General Procedure for the Synthesis of 1-Thio-β-D-glucopyranoside Tetraacetates (Adapted from )
α-D-Glucose pentaacetate (0.5–2.1 g and 1.3–5.3 mmol) was dissolved in CH2Cl2 (25 mL), followed by addition of a thiol (1.5 mol eq). The reaction mixture was stirred for 10 minutes in an ice bath. Then BF3Et2O (3 mol eq) was added slowly, and the mixture was allowed to come to room temperature and was stirred with gentle reflux overnight. The solvent was removed by evaporation and product was isolated by column chromatography (silica gel, hexane/EtOAc 5 : 1).
3.1.1. Phenyl-1-thio-β-D-glucopyranoside Tetraacetate (1a)
The stirring overnight was done without reflux. Yield 1.01 g (45%) from 2.00 g (5.1 mmol) α-D-glucose pentaacetate. 1H NMR (CDCl3): δ 2.00 (s, CH3), 2.02 (s, CH3), 2.087 (s, CH3), 2.094 (s, CH3), 3.73 (ddd, J = 2.5, 5.1, 10.0 Hz; H5), 4.18 (dd, J = 2.5, 12.2 Hz; H6), 4.23 (dd, J = 5.1, 12.2 Hz; H6), 4.71 (d, J = 10.1 Hz; H1), 4.98 (dd, J = 9.3, 10.1 Hz; H2), 5.05 (dd, J = 9.6, 10.0 Hz; H4), 5.23 (t, J = 9.4 Hz; H3), 7.31–7.35 (m, 3H; Ph), 7.50 (m, 2H; Ph). 13C NMR (CDCl3): 20.60, 20.75 (CH3, Ac), 62.09 (C6), 68.10 (C4), 69.86 (C2), 73.91 (C3), 75.74 (C5), 85.71 (C1), 128.41, 128.92, 131.59, 133.10 (Ph), 169.26, 169.40, 170.18, 170.58 (C=O); (Lit. 1H, 13C NMR ). HRMS: C20H24O9S requires m/z [M+NH4]+ 458.1485; observed m/z 458.1468.
3.1.2. p-Chlorophenyl-1-thio-β-D-glucopyranoside Tetraacetate (1b)
Yield 0.97 g (39%) from 2.07 g (5.3 mmol) α-D-glucose pentaacetate. 1H NMR (CDCl3): δ 1.99 (s, CH3), 2.02 (s, CH3), 2.09 (s, CH3), 2.10 (s, CH3), 3.72 (ddd, J = 2.6, 4.9, 10.1 Hz; H5), 4.18 (dd, J = 2.6, 12.3 Hz; H6), 4.22 (dd, J = 4.9, 12.3 Hz; H6), 4.65 (d, J = 10.1 Hz; H1), 4.94 (dd, J = 9.4, 10.1 Hz; H2), 5.02 (t, J = 9.8 Hz; H4), 5.22 (t, J = 9.4 Hz; H3), 7.30 (m, 2H; Ar), 7.44 (m, 2H; Ar). 13C NMR (CDCl3): 20.56, 20.72 (CH3, Ac), 61.96 (C6), 67.99 (C4), 69.71 (C2), 73.79 (C3), 75.81 (C5), 85.16 (C1), 129.04, 129.38, 134.97 (Ar), 169.21, 169.35, 170.13, 170.51 (C=O); (Lit. 1H, 13C NMR ). HRMS: C20H23ClO9S requires m/z [M+NH4]+ 492.1095; observed m/z 492.1062.
3.1.3. p-Tolyl-1-thio-β-D-glucopyranoside Tetraacetate (1c)
Yield 1.75 g (71%) from 2.05 g (5.3 mmol) α-D-glucose pentaacetate. 1H NMR (CDCl3): δ 1.99 (s, CH3), 2.02 (s, CH3), 2.09 (s, CH3), 2.10 (s, CH3), 2.35 (s, CH3, p-tolyl), 3.70 (ddd, J = 2.6, 4.9, 10.1 Hz; H5), 4.18 (dd, J = 2.6, 12.2 Hz; H6), 4.22 (dd, J = 4.9, 12.2 Hz; H6), 4.64 (d, J = 10.1 Hz; H1), 4.94 (dd, J = 9.4, 10.0 Hz; H2), 5.03 (t, J = 9.8 Hz; H4), 5.21 (t, J = 9.4 Hz; H3), 7.13 (d, J = 7.8 Hz, 2H; Ar), 7.39 (d, J = 8.0 Hz, 2H; Ar). 13C NMR (CDCl3): 20.68, 20.70, 20.85, 20.87 (CH3, Ac), 21.29 (CH3, p-tolyl), 62.08 (C6), 68.12 (C4), 69.85 (C2), 73.97 (C3), 75.69 (C5), 85.79 (C1), 127.49, 129.65, 133.81, 138.79 (Ar), 169.23, 169.38, 170.19, 170.58 (C=O); (Lit. 1H, 13C NMR ). HRMS: C21H26O9S requires m/z [M+NH4]+ 472.1641; observed m/z 472.1590.
3.1.4. (2-Naphthyl)-1-thio-β-D-glucopyranoside Tetraacetate (1d)
Yield 0.23 g (37%) from 0.50 g (1.3 mmol) α-D-glucose pentaacetate dissolved in 10 mL CH2Cl2. 1H NMR (CDCl3): δ 1.99 (s, CH3), 2.02 (s, CH3), 2.04 (s, CH3), 2.12 (s, CH3), 3.74 (ddd, J = 2.4, 5.1, 10.1 Hz; H5), 4.18 (dd, J = 2.4, 12.2 Hz; H6), 4.24 (dd, J = 5.2, 12.3 Hz; H6), 4.79 (d, J = 10.1 Hz; H1), 5.02 (dd, J = 9.3, 10.0 Hz; H2), 5.05 (dd, J = 9.5, 10.0 Hz; H4), 5.24 (t, J = 9.4 Hz; H3), 7.51 (m, 2H; Ar), 7.56 (dd, J = 1.8, 8.6 Hz, 1H; Ar), 7.79 (d, J = 8.5 Hz, 1H; Ar), 7.82 (m, 2H; Ar), 8.00 (d, J = 1.7 Hz, 1H; Ar). 13C NMR (CDCl3): 20.58, 20.70, 20.79 (CH3, Ac), 62.10 (C6), 68.14 (C4), 69.98 (C2), 73.92 (C3), 75.82 (C5), 85.80 (C1), 126.65, 126.75, 127.68, 128.46, 128.70, 130.19, 132.74, 132.84, 133.39 (Ar), 169.29, 169.36, 170.18, 170.59 (C=O); (Lit. 1H, 13C NMR ). HRMS: C24H26O9S requires m/z [M+NH4]+ 508.1641; observed m/z 508.1583.
3.1.5. Benzyl-1-thio-β-D-glucopyranoside Tetraacetate (1e)
Yield 1.09 g (46%) from 2.05 g (5.3 mmol) α-D-glucose pentaacetate. 1H NMR (CDCl3): δ 1.99 (s, CH3), 2.01 (s, CH3), 2.02 (s, CH3), 2.12 (s, CH3), 3.57 (ddd, J = 2.3, 5.1, 10.0 Hz; H5), 3.81 (d, J = 12.9 Hz, 1H; CH2Ph), 3.91 (d, J = 12.9 Hz, 1H; CH2Ph) 4.11 (dd, J = 2.3, 12.4 Hz; H6), 4.21 (dd, J = 5.1, 12.4 Hz; H6), 4.26 (d, J = 10.0 Hz; H1), 5.04 (dd, J = 9.3, 10.0 Hz; H2), 5.05 (dd, J = 9.3, 10.0 Hz; H4), 5.11 (t, J = 9.3 Hz; H3), 7.27–7.34 (m, 5H; Ph). 13C NMR (CDCl3): 20.56, 20.58, 20.64, 20.76 (CH3, Ac), 33.77 (CH2Ph), 62.18 (C6), 68.31 (C4), 69.74 (C2), 73.78 (C3), 75.76 (C5), 81.89 (C1), 127.41, 128.59, 129.04, 136.76 (Ph), 169.40, 170.18, 170.63 (C=O); (Lit. 1H, 13C NMR ). HRMS: C21H26O9S requires m/z [M+NH4]+ 472.1641; observed m/z 472.1596.
3.2. General Procedure for the Synthesis of 1-Thio-β-D-glucopyranosides (Adapted from )
The respective 1-thio-β-D-glucopyranoside tetraacetate 1a–1e (100 mg) was mixed with MeOH (2 mL) in a 5 mL flask. The freshly prepared MeONa solution was added slowly until the pH of the mixture was 9-10 as measured by pH paper. The reaction mixture was allowed to stir until all starting tetraacetate was consumed (TLC; silica gel, hexane/EtOAc 5 : 1). The basic solution was then neutralized with Dowex 50W X8 ion exchange resin, filtered, and evaporated to yield 95–99% of pure thioglucoside.
3.2.1. Phenyl-1-thio-β-D-glucopyranoside (2a)
100 mg (0.227 mmol) 1a yielded 60 mg (97%) 2a as a white solid, mp 131–133°C (Lit. 113°C ). 1H NMR (CD3OD): δ 3.21 (dd, J = 8.8, 9.8 Hz; H2), 3.28 (m, J = 8.5 Hz; H3), 3.32 (ddd, J = 2.3, 5.7, 9.7 Hz; H5), 3.38 (t, J = 8.6 Hz; H4), 3.66 (dd, J = 5.5, 12.1 Hz; H6), 3.86 (dd, J = 2.1, 12.1 Hz; H6), 4.59 (d, J = 9.8 Hz; H1), 7.22 (m, 1H; Ph), 7.29 (m, 2H; Ph), 7.55 (m, 2H; Ph). 13C NMR (CD3OD): 62.85 (C6), 71.32 (C4), 73.75 (C2), 79.68 (C3), 82.04 (C5), 89.41 (C1), 128.29, 129.87, 132.66, 135.32 (Ph); (Lit. 1H, 13C NMR ). HRMS: C12H16O5S requires m/z [M+NH4]+ 290.1062; observed m/z 290.0993.
3.2.2. p-Chlorophenyl-1-thio-β-D-glucopyranoside (2b)
100 mg (0.21 mmol) 1b yielded 61 mg (94%) 2b as a white solid, mp 169.5–171.5°C (Lit. 172–175°C ). 1H NMR (CD3OD): δ 3.16 (dd, J = 8.8, 9.7 Hz; H2), 3.25 (dd, J = 8.9, 9.6 Hz; H3), 3.32 (ddd, J = 2.2, 5.7, 9.7 Hz; H5), 3.35 (t, J = 8.7 Hz; H4), 3.63 (dd, J = 5.7, 12.1 Hz; H6), 3.85 (dd, J = 2.1, 12.1 Hz; H6), 4.55 (d, J = 9.8 Hz; H1), 7.28 (m, 2H; Ar), 7.52 (m, 2H; Ar). 13C NMR (CD3OD): 62.81 (C6), 71.29(C4), 73.67 (C2), 79.63 (C3), 82.10 (C5), 89.11 (C1), 129.88, 134.04, 134.28, 134.41 (C, Ar). HRMS: C12H15ClO5S requires m/z [M+NH4]+ 324.0673; observed m/z 324.0619.
3.2.3. p-Tolyl-1-thio-β-D-glucopyranoside (2c)
100 mg (0.22 mmol) 1c yielded 62 mg (98%) 2c as a white solid, mp 139.5–142°C (Lit. 148°C ). 1H NMR (CD3OD): δ 2.30 (s, CH3), 3.16 (dd, J = 8.8, 9.7 Hz; H2), 3.25 (t, J = 9.8 Hz; H3), 3.28 (ddd, J = 2.1, 5.5, 9.6 Hz; H5), 3.35 (t, J = 8.6 Hz; H4), 3.64 (dd, J = 5.4, 12.1 Hz; H6), 3.84 (dd, J = 1.9, 12.0 Hz; H6), 4.50 (d, J = 9.8 Hz; H1), 7.11 (m, 2H; Ar), 7.45 (m, 2H; Ar). 13C NMR (CD3OD): 21.10 (CH3), 62.87 (C6), 71.35 (C4), 73.70 (C2), 79.68 (C3), 81.96 (C5), 89.64 (C1), 130.53, 131.21, 133.48, 138.74 (C, Ar); (Lit. 1H, 13C NMR ). HRMS: C13H18O5S requires m/z [M+NH4]+ 304.1219; observed m/z 304.1214.
3.2.4. (2-Naphthyl)-1-thio-β-D-glucopyranoside (2d)
100 mg (0.20 mmol) 1d yielded 62 mg (94%) 2d as a white solid, mp 122–123.5°C (Lit. 117–125°C ). 1H NMR (CD3OD): δ 3.26 (dd, J = 9.8, 8.8 Hz; H2), 3.29 (m, H3), 3.32 (ddd, J = 2.3, 6.0, 9.8 Hz; H5), 3.38 (t, J = 8.9 Hz; H4), 3.66 (dd, J = 6.0, 12.1 Hz; H6), 3.86 (dd, J = 2.2, 12.1 Hz; H6), 4.71 (d, J = 9.8 Hz; H1), 7.45 (m, 2H; Ar), 7.63 (m, 1H; Ar), 7.79 (m, 3H; Ar), 8.07 (m, 1H; Ar). 13C NMR (CD3OD): 62.89 (C6), 71.39 (C4), 73.82 (C2), 79.70 (C3), 82.16 (C5), 89.37 (C1), 127.18, 127.52, 128.53, 128.65, 129.23, 130.13, 131.16, 132.75, 133.81, 135.14 (Ar). HRMS: C16H18O5S requires m/z [M+NH4]+ 340.1219; observed m/z 340.1218.
3.2.5. Benzyl-1-thio-β-D-glucopyranoside (2e)
100 mg (0.22 mmol) 1c yielded 61 mg (97%) 2c as a clear oil. 1H NMR (CD3OD): δ 3.19–3.29 (m, 4H; H2+H3+H4+H5), 3.67 (dd, J = 6.2, 12.1 Hz; H6), 3.85 (d, J = 12.7 Hz, 1H; CH2Ph), 3.89 (dd, J = 2.2, 12.1 Hz; H6), 4.03 (d, J = 12.7 Hz, 1H; CH2Ph), 4.17 (d, J = 9.3 Hz; H1), 7.22 (t, J = 7.4 Hz, 1H; Ph), 7.29 (t, J = 7.3 Hz, 2H; Ph), 7.37 (m, 2H; Ph). 13C NMR (CD3OD): 32.98 (CH2Ph), 61.67 (C6), 70.27 (C4), 73.02 (C2), 78.35(C3), 80.68 (C5), 83.78 (C1), 126.66, 128.11, 128.97, 138.10 (Ph); (Lit. 1H, 13C NMR ). HRMS: C13H18O5S requires m/z [M+NH4]+ 304.1219; observed m/z 304.1202.
3.3. Hexyl-1-thio-β-D-glucopyranoside (2f)
1-Thio-β-D-glucopyranose sodium salt 3 (100 mg, 0.46 mmol) was mixed with methanol (2 mL) followed by the addition of bromohexane (75 μL, 0.6 mmol). The reaction mixture was stirred at room temperature overnight and then evaporated in vacuo, and the resulting white paste was rinsed with hexane and then redissolved in methanol. The resulting suspension was filtered, and the filtrate evaporated to yield clear oil 2f (98 mg, 76%). 1H NMR (CD3OD): δ 0.90 (t, J = 11.4 Hz, 3H; CH3) 1.31 (m, 4H; CH2), 1.40 (m, 2H; CH2), 1.62 (m, 2H; CH2), 2.68 (dt, J = 12.5, 7.5 Hz, 1H; SCH2), 2.73 (dt, J = 12.4, 7.3 Hz, 1H; SCH2), 3.18 (dd, J = 8.7, 9.7 Hz; H2), 3.25 (ddd, J = 2.1, 5.5, 9.5 Hz; H5), 3.29 (m, H3), 3.33 (t, J = 8.9 Hz; H4), 3.64 (dd, J = 5.5, 12.0 Hz; H6), 3.84 (dd, J = 2.2, 12.0 Hz; H6), 4.33 (d, J = 9.8 Hz; H1). 13C NMR (CD3OD): 13.05 (CH3), 22.30 (CH2), 28.36 (CH2), 29.52 (CH2), 29.68 (CH2), 31.26 (CH2), 61.58 (C6), 70.17 (C4), 73.06 (C2), 78.27 (C3), 80.79 (C5), 85.81 (C1); (Lit. 1H, 13C NMR ). HRMS: C12H24O5S requires m/z [M+NH4]+ 298.1688; observed m/z 298.1661.
3.4. (1,10-Phenanthrolin-5-yl)-1-thio-β-D-glucopyranoside (2g) 
1-Thio-β-D-glucopyranose sodium salt 3 (65 mg, 0.30 mmol) in dry ethanol (2 mL) was added to a solution of 5,6-epoxy-5,6-dihydro-1,10-phenanthroline (50 mg, 0.255 mmol) in dry ethanol (2 mL) and stirred at room temperature for 36 h until complete conversion of the epoxide as monitored by TLC (silica gel, EtOAc/CH3OH/NH4OH 7 : 3 : 1). The precipitate was collected and recrystalized from EtOH affording a yellowish solid product (61 mg, 64%): mp 154-155°C; −64.0° (c 0.38, DMSO). 1H NMR (DMSO): 3.08 (t, J = 9.2 Hz; H4), 3.17 (dd, J = 9.6, 8.8 Hz; H2), 3.23 (t, J = 8.7 Hz; H3), 3.30 (ddd, J = 9.6, 6.5, 2.0 Hz; H5), 3.41 (dd, J = 11.8, 6.4 Hz; H6), 3.69 (dd, J = 11.8, 1.9 Hz; H6), 4.79 (d, J = 9.7 Hz; H1), 7.74 (dd, J = 8.1, 4.3 Hz; H8′), 7.80 (dd, J = 8.3, 4.2 Hz; H3′), 8.29 (s, H6′), 8.36 (dd, J = 8.1, 1.6 Hz; H4′), 8.79 (dd, J = 8.4, 1.6 Hz; H7′), 9.02 (dd, J = 4.3, 1.6 Hz; H9′), 9.09 (dd, J = 4.2, 1.6 Hz; H2′). 13C NMR (DMSO): 61.57 (C6), 70.36 (C4), 73.17 (C2), 79.74 (C3), 81.75 (C5), 87.46 (C1), 123.91 (C3′), 124.29 (C8′), 128.73, 128.82 (C4a′/C6a′), 129.13 (C6′), 131.44 (C5′), 134.13 (C4′), 136.34 (C7′), 145.20, 145.86 (C10a′/C10b′), 150.48, 150.55 (C2′/C9′). HRMS: C30H26N4O2S2 requires m/z [M+H]+ 375.1015; observed m/z 375.1022.
3.5. Diethyl 7-oxabicyclo[4.1.0]heptane-trans-3,4-dicarboxylate (4)
The epoxide 4 was prepared as described before  and isolated by column chromatography (Al2O3, gradient washing with hexane/EtOAc 8 : 2 → 7 : 3) as a colorless liquid (60%). 1H NMR (CDCl3): δ 1.21 (t, J = 7.1 Hz, 3H; CH3), 1.22 (t, J = 7.1 Hz, 3H; CH3), 1.87 (ddd, J = 14.9, 10.8, 2.1 Hz; H2), 2.04 (dd, J = 15.5, 10.9 Hz; H5), 2.30 (ddd, J = 15.5, 6.6, 4.8 Hz; H5), 2.45 (ddd, J = 14.9, 4.8, 1.8 Hz; H2), 2.58 (dt, J = 10.7, 6.7 Hz; H4), 2.80 (dt, J = 10.7, 4.9 Hz; H3), 3.17 (t, J = 4.3 Hz; H6), 3.24 (m, H1), 4.11 (m, 4H; OCH2). 13C NMR (CDCl3): δ 14.20 (CH3), 26.44 (C5), 27.30 (C2), 37.80 (C3), 40.14 (C4), 50.40 (C6), 52.00 (C1), 60.84 (OCH2), 173.76, 174.82 (C=O). HRMS: C12H18O5 requires [M+H]+ m/z 243.1233; observed m/z 243.1240.
3.6. [(1S*,2S*,4S*,5S*)-2-Hydroxy-4,5-bis(ethoxycarbonyl)-cyclohexyl]-1-thio-β-D-glucopyranoside (2h)
1-Thio-β-D-glucopyranose sodium salt 3 (100 mg, 0.46 mmol) in 7 mL of ethanol was added to a suspension of epoxide 4 (80 mg, 0.33 mmol) in ethanol (2 mL) at room temperature. Reaction mixture was stirred at 40°C for 12 h until complete conversion of the epoxide as monitored by TLC (silica gel, EtOAc/MeOH 4 : 1). The solvent was evaporated under vacuum and residue was purified by column chromatography (silica gel, EtOAc/MeOH 7 : 3) affording product 2h (a mixture of diastereomers 1 : 1) as a white solid (55 mg, 38%). 1H NMR (CD3OD): δ 1.21 (t, J = 7.1 Hz, 6H; 2CH3), 1.89 (dd, J = 8.3, 3.4 Hz, 1H; H3′); 1.94−2.05 (m, 2H; H3′+H6′), 2.21, 2.24 (dt, J = 3.9, 11.6 Hz, 1H; H6′), 2.86, 2.89 (dt, J = 3.7, 11.3 Hz; H4′), 2.98 (m, H5′), 3.17 (t, J = 8.6 Hz; H2), 3.23–3.35 (m, 4H; H1′+H3+H4+H5), 3.62, 3.65 (dd, J = 12.1, 5.1 Hz, 1H; H6), 3.84 (dd, J = 12.1, 1.8 Hz, 1H; H6), 3.98, 4.02 (br dd, J = 6.4, 3.3 Hz; H2′), 4.06–4.14 (m, 4H; OCH2), 4.41, 4.42 (d, J = 9.8 Hz; H1). 13C NMR (CD3OD): δ 13.14 (CH3), 28.51, 28.92 (C6′), 30.29, 30.45 (C3′), 39.09, 39.14 (C5′), 40.22, 40.24 (C4′), 44.34, 44.89 (C1′), 60.53, 60.58 (OCH2), 61.53 (C6), 67.73, 68.36 (C2′), 69.92, 70.01 (C4), 73.12, 73.18 (C2), 78.21 (C3), 80.77 (C5), 85.91, 86.00 (C1), 174.88, 175.18 (C=O). HRMS: C18H30O10S requires m/z [M+NH4]+ 456.1903, [M+H]+ 439.1638; observed m/z 456.1967, 439.1654.
3.7. [(1S*,2S*,4S*,5S*)-2-Hydroxy-4,5-bis (hydroxycarbonyl)-cyclohexyl]-1-thio-β-D-glucopyranoside (2i)
1 mL of KOH (1 M) was added to a solution of diester 2h (150 mg, 0.34 mmol) in 2 mL of methanol and mixture stirred for 72 h at room temperature. HCl (1 M) was added to reaction mixture till pH 6 and solvent was evaporated under vacuum. Residue was purified by column chromatography (silica gel, EtOAc/CH3OH 1 : 4) affording product 2i (one diastereomer) as a white solid (40 mg, 31%; mp > 240°C). 1H NMR (CD3OD): δ 1.85 (dd, J = 13.4, 3.8 Hz; H3′), 1.93–2.05 (m, 2H; H6′+H3′), 2.26 (m, 1H; H6′), 2.85 (m, H5′), 2.95 (m, H4′), 3.19 (t, J = 9.1 Hz; H2), 3.21–3.35 (m, 4H; H1′+H3+H4+H5), 3.64 (m, 1H; H6), 3.84 (m, 1H; H6), 3.95 (m, H2′), 4.43 (d, J = 9.8 Hz; H1). 13C NMR (CD3OD): δ 29.23 (C6′), 29.66 (C3′), 31.02 (C4′), 31.18 (C5′), 48.53 (C1′), 61.46 (C6), 69.05 (C2′), 69.98 (C4), 73.17 (C2), 78.24, 80.72 (C3/C5), 85.99 (C1), 179.73 (C=O). HRMS: C14H22O10S requires m/z [M+NH4]+ 400.1277; observed m/z 400.1289.
3.8. Procedures for the Glycosidase Inhibition and Activation Assay
The enzyme activities (α/β-D-galactosidases and α/β-D-glucosidases) were assayed using multienzyme complexes isolated from fungi P. canescens and A. oryzae as described before [25–27]. All assays were performed in a standard way by monitoring spectrophotometrically (with Beckman Du-65 spectrometer) the release of p-nitrophenol from the corresponding p-nitrophenyl glycosides at 30°C. One unit of enzyme activity was defined as the amount of enzyme that releases 1 μmol of p-nitrophenol per minute. Enzyme and substrate concentrations were selected so that the degree of hydrolysis was never more than 20% and in most cases was less than 10%, over the course of the assay. The method used to measure the rate of the reaction assumes that the amount of the substrate is high enough, such that the disappearance over a given period is insignificant; that is, the rate of the reaction is close to linear for the first stage of the reaction.
For the preliminary estimation, the enzyme solutions (100 μL with activities μU) were mixed with a set of inhibitor/activator solutions (100 μL, 10 mM) and then diluted with 700 μL of 0.2 M acetic buffer (pH 4.2), and the mixture was incubated for 1 h at 30°C. The reaction was initiated with addition of a proper substrate (100 μL of 20 mM p-nitrophenyl glycopyranoside), and aliquots were taken after 5 and 10 min. The reaction was terminated by addition of 1 mL of 1 M Na2CO3 to 0.5 mL of aliquot solution. The concentration of the released p-nitrophenol was determined at 400 nm using molar extinction coefficient 18.3 mM−1 cm−1. The inhibition/activation was estimated as a loss/increase of enzymatic activity in % (Figures 1 and 2).
The same protocol was used for estimation of the kinetic parameters with the following changes: 1 mL of enzyme solution, 0.5 mL inhibitor solution, and 3.0 mL of 0.2 M acetic buffer (pH 4.2) were incubated for 1 h at 30°C before addition of 0.5 mL of substrate (20 mM). Six substrate solutions were prepared with various concentrations up to saturation point for each enzyme. The final substrate concentration after all additions varied from 0.01 mM to 2.00 mM. The final inhibitor concentrations were 330 μM for A. oryzae and 33 μM for P. canescens. Reaction rates were measured as described above. Control experiments with no enzyme were performed to exclude the errors due to the substrate spontaneous hydrolysis. Enzyme behavior (at pH 4.2 and 30°C) abided the Michaelis-Menten equation. Parameters and were calculated from Lineweaver-Burk (double-reciprocal) plot [52, 53]. The values were used to calculate by the following equation: Three independent trials using freshly prepared substrate and enzyme solutions were performed to obtain each parameter. Values for were reproducible within ±5% and ±10%, respectively, and the standard error was ±15% for the values (Table 1).
The results of our studies show that simple and readily available 1-thio-β-D-glucopyranosides can be potent inhibitors for α- and β-D-glucosidases. The efficiency of inhibition strongly depends on the structure of aglycon and generally increases for β-glucosidases with increase of its lipophilicity. Another interesting feature of the studied compounds is an activation of α- and β-galactosidases.
Conflict of Interests
The authors declare that there is no conflict of interests regarding the publication of this paper.
The funding of the JEOL ECA-600 NMR spectrometer by the NSF Instrumentation Grant (NSF-MRI-0722654) and the support from the Department of Chemistry, UOP, are gratefully acknowledged. Irina A. Dotsenko appreciates the John P. Shinkai Graduate Students Scholarship. The authors thank Professor O. David Sparkman and Dr. Matthew Curtis for the use of Pacific Mass Spectrometry Facility.
- R. K. H. Kinne and F. Castaneda, “SGLT inhibitors as new therapeutic tools in the treatment of diabetes,” Handbook of Experimental Pharmacology, vol. 203, pp. 105–126, 2011.
- A. Dondoni and A. Marra, “Calixarene and calixresorcarene glycosides: their synthesis and biological applications,” Chemical Reviews, vol. 110, no. 9, pp. 4949–4977, 2010.
- F. Castaneda, A. Burse, W. Boland, and R. K. H. Kinne, “Thioglycosides as inhibitors of hSGLT1 and hSGLT2: potential therapeutic agents for the control of hyperglycemia in diabetes,” International Journal of Medical Sciences, vol. 4, no. 3, pp. 131–139, 2007.
- Q. V. Vo, C. Trenerry, S. Rochfort, J. Wadeson, C. Leyton, and A. B. Hughes, “Synthesis and anti-inflammatory activity of aromatic glucosinolates,” Bioorganic and Medicinal Chemistry, vol. 21, no. 19, pp. 5945–5954, 2013.
- J. Rodrigue, G. Ganne, B. Blanchard et al., “Aromatic thioglycoside inhibitors against the virulence factor LecA from Pseudomonas aeruginosa,” Organic and Biomolecular Chemistry, vol. 11, no. 40, pp. 6906–6918, 2013.
- E. S. H. El Ashry, E. S. H. El Tamany, M. E. D. A. El Fattah, A. T. A. Boraei, and H. M. Abd El-Nabi, “Regioselective synthesis, characterization and antimicrobial evaluation of S-glycosides and S,N-diglycosides of 1,2-Dihydro-5-(1H-indol-2-yl)-1,2,4-triazole-3-thione,” European Journal of Medicinal Chemistry, vol. 66, pp. 106–113, 2013.
- H. A. El-Sayed, A. H. Moustafa, and A. E. Z. Haikal, “Synthesis, antiviral, and antimicrobial activity of 1,2,4-triazole thioglycoside derivatives,” Phosphorus, Sulfur and Silicon and the Related Elements, vol. 188, no. 5, pp. 649–662, 2013.
- A. J. Cagnoni, O. Varela, J. Kovensky, and M. L. Uhrig, “Synthesis and biological activity of divalent ligands based on 3-deoxy-4-thiolactose, an isosteric analogue of lactose,” Organic and Biomolecular Chemistry, vol. 11, no. 33, pp. 5500–5511, 2013.
- E. Repetto, C. Marino, and O. Varela, “Synthesis of the (1→6)-linked thiodisaccharide of galactofuranose: inhibitory activity against a β-galactofuranosidase,” Bioorganic and Medicinal Chemistry, vol. 21, no. 11, pp. 3327–3333, 2013.
- I. L. Rogers, D. W. Gammon, and K. J. Naidoo, “Conformational preferences of plumbagin with phenyl-1-thioglucoside conjugates in solution and bound to MshB determined by aromatic association,” Carbohydrate Research, vol. 371, pp. 52–60, 2013.
- S. V. Pestova, D. V. Sudarikov, S. A. Rubtsova, and A. V. Kutchin, “Synthesis and asymmetric oxidation of thioglycosides derived from neomenthanethiol and α-D-galactose,” Russian Journal of Organic Chemistry, vol. 49, no. 3, pp. 366–373, 2013.
- T. Terauchi, Y. Koyama, S. Machida, T. Kasumi, and S. Komba, “Synthesis of novel thioglycoside analogs as the substrates and/or the inhibitors of cellobiohydrolases,” Journal of Applied Glycoscience, vol. 59, no. 1, pp. 11–19, 2012.
- C. Stanetty, A. Wolkerstorfer, H. Amer et al., “Synthesis and antiviral activities of spacer-linked 1-thioglucuronide analogs of glycyrrhizin,” Beilstein Journal of Organic Chemistry, vol. 8, pp. 705–711, 2012.
- R. Dettmann and T. Ziegler, “Synthesis of octyl S-glycosides of tri- to pentasaccharide fragments related to the GPI anchor of Trypanosoma brucei,” Carbohydrate Research, vol. 346, no. 15, pp. 2348–2361, 2011.
- H. N. Hafez, H. A. R. Hussein, and A.-R. B. A. El-Gazzar, “Synthesis of substituted thieno[2,3-d]pyrimidine-2,4-dithiones and their S-glycoside analogues as potential antiviral and antibacterial agents,” European Journal of Medicinal Chemistry, vol. 45, no. 9, pp. 4026–4034, 2010.
- M. Poláková, M. Beláňová, L. Petruš, and K. Mikušová, “Synthesis of alkyl and cycloalkyl α-D-mannopyranosides and derivatives thereof and their evaluation in the mycobacterial mannosyltransferase assay,” Carbohydrate Research, vol. 345, no. 10, pp. 1339–1347, 2010.
- R. Caraballo, M. Sakulsombat, and O. Ramström, “Towards dynamic drug design: identification and optimization of β-galactosidase inhibitors from a dynamic hemithioacetal system,” ChemBioChem, vol. 11, no. 11, pp. 1600–1606, 2010.
- I. F. Nassar, “Synthesis and antitumor activity of new substituted mercapto-1,2,4-triazine derivatives, their thioglycosides, and acyclic thioglycoside analogs,” Journal of Heterocyclic Chemistry, vol. 50, no. 1, pp. 129–134, 2013.
- I. García-Álvarez, H. Groult, J. Casas et al., “Synthesis of antimitotic thioglycosides: in vitro and in vivo evaluation of their anticancer activity,” Journal of Medicinal Chemistry, vol. 54, no. 19, pp. 6949–6955, 2011.
- M. A. Abu-Zaied, E. M. El-Telbani, G. H. Elgemeie, and G. A. M. Nawwar, “Synthesis and in vitro anti-tumor activity of new oxadiazole thioglycosides,” European Journal of Medicinal Chemistry, vol. 46, no. 1, pp. 229–235, 2011.
- M. A. Jermyn, “Fungal cellulases. XV. Acceptor specificity of the aryl beta-glucosidase of Stachybotrys atra.,” Australian Journal of Biological Sciences, vol. 19, no. 5, pp. 903–917, 1966.
- T. Kuntothom, M. Raab, I. Tvaroška et al., “Binding of β-d-glucosides and β-d-mannosides by rice and barley β-d-glycosidases with distinct substrate specificities,” Biochemistry, vol. 49, no. 40, pp. 8779–8793, 2010.
- M. Schnabelrauch, A. Vasella, and S. G. Withers, “Synthesis and evaluation as irreversible glycosidase inhibitors of mono- and oligo(glycosylthio)benzoquinones,” Helvetica Chimica Acta, vol. 77, no. 3, pp. 778–799, 1994.
- C.-S. Kuhn, J. Lehmann, and J. Steck, “Syntheses and properties of some photolabile β-thioglycosides: potential photoaffinity reagents for β-glycoside hydrolases,” Tetrahedron, vol. 46, no. 9, pp. 3129–3134, 1990.
- B. Brazdova, N. S. Tan, N. M. Samoshina, and V. V. Samoshin, “Novel easily accessible glucosidase inhibitors: 4-hydroxy-5-alkoxy-1,2-cyclohexanedicarboxylic acids,” Carbohydrate Research, vol. 344, no. 3, pp. 311–321, 2009.
- N. S. Tan, B. Brazdova, N. M. Samoshina, and V. V. Samoshin, “Novel inhibitors for fungal glycosidases based on cyclohexane-1, 2-dicarboxylic acids,” Journal of Undergraduate Chemistry Research, vol. 6, no. 4, pp. 186–192, 2007.
- I. A. Dotsenko, M. Curtis, N. M. Samoshina, and V. V. Samoshin, “Convenient synthesis of 5-aryl(alkyl)sulfanyl-1,10-phenanthrolines from 5,6-epoxy-5,6-dihydro-1,10-phenanthroline, and their activity towards fungal β-d-glycosidases,” Tetrahedron, vol. 67, no. 39, pp. 7470–7478, 2011.
- M. M. Gomarteli, A. K. Kulikova, A. K. Tsereteli, A. M. Bezborodov, and G. I. Kvesitadze, “beta-Galactosidase from Penicillium canescens st. 20171,” Applied Biochemistry and Microbiology, vol. 24, no. 1, pp. 16–22, 1988.
- N. M. Samoshina, L. V. Yugova, P. H. Gross, G. B. Bravova, A. A. Shishkova, and V. V. Samoshin, “Partial purification and characterization of glycosidases from Aspergillus oryzae and Penicillium canescens,” in Proceedings of the 219th National Meeting of the American Chemical Society, p. 160, The American Chemical Society, San Francisco, Calif, USA, 2000.
- O. S. Korneeva, N. A. Zherebtsov, and I. V. Cheryomushkina, “Identification of catalytically active groups of Penicillium canescens F-436 beta-galactosidase,” Biochemistry, vol. 66, no. 3, pp. 334–339, 2001.
- O. A. Sinitsyna, F. E. Bukhtoyarov, A. V. Gusakov et al., “Isolation and properties of major components of Penicillium canescens extracellular enzyme complex,” Biochemistry, vol. 68, no. 11, pp. 1200–1209, 2003.
- I. M. Gracheva and A. Y. Krivova, The Technology of Enzyme Preparations, Elevar, Moscow, Russia, 2000.
- A. B. Pardee, F. Jacob, and J. Monod, “The genetic control and cytoplasmic expression of inducibility in the synthesis of β-galactosidase by Escherichia coli,” Journal of Molecular Biology, vol. 1, pp. 165–178, 1959.
- C. Riou, J. Salmon, M. Vallier, Z. Günata, and P. Barre, “Purification, characterization, and substrate specificity of a novel highly glucose-tolerant β-glucosidase from Aspergillus oryzae,” Applied and Environmental Microbiology, vol. 64, no. 10, pp. 3607–3614, 1998.
- X. Bian, X. Fan, C. Ke, Y. Luan, G. Zhao, and A. Zeng, “Synthesis and α-glucosidase inhibitory activity evaluation of N-substituted aminomethyl-β-d-glucopyranosides,” Bioorganic and Medicinal Chemistry, vol. 21, no. 17, pp. 5442–5450, 2013.
- K. Duskova, L. Gude, and M.-S. Arias-Pérez, “N-Phenanthroline glycosylamines: synthesis and copper(II) complexes,” Tetrahedron, vol. 70, no. 5, pp. 1071–1076, 2014.
- T. M. Wrodnigg and A. E. Stütz, “The two faces of iminoalditols: powerful inhibitors trigger glycosidase activation,” Current Enzyme Inhibition, vol. 8, no. 1, pp. 47–99, 2012.
- Y. Nakagawa, S. Sehata, S. Fujii, H. Yamamoto, A. Tsuda, and K. Koumoto, “Mechanistic study on the facilitation of enzymatic hydrolysis by α-glucosidase in the presence of betaine-type metabolite analogs,” Tetrahedron, vol. 70, no. 35, pp. 5895–5903, 2014.
- N. M. Samoshina and V. V. Samoshin, “The Michaelis constants ratio for two substrates with a series of fungal (mould and yeast) β-galactosidases,” Enzyme and Microbial Technology, vol. 36, no. 2-3, pp. 239–251, 2005.
- E. Alverson-Banks Avegno, S. J. Hasty, A. R. Parameswar, G. S. Howarth, A. V. Demchenko, and L. D. Byers, “Reactive thioglucoside substrates for β-glucosidase,” Archives of Biochemistry and Biophysics, vol. 537, no. 1, pp. 1–4, 2013.
- A. E. Stutz, Ed., Iminosugars as Glycosidase Inhibitors: Nojirimycin and Beyond, Wiley-VCH, Weinheim, Germany, 1999.
- P. Compain and O. R. Martin, Eds., Iminosugars: From Synthesis to Therapeutic Applications, John Wiley & Sons, Chichester, UK, 2007.
- P. Compain, V. Chagnault, and O. R. Martin, “Tactics and strategies for the synthesis of iminosugar C-glycosides: a review,” Tetrahedron Asymmetry, vol. 20, no. 6-8, pp. 672–711, 2009.
- G. M. Aerts, O. van Opstal, and C. K. de Bruyne, “Mixed inhibition of β-d-glucosidase from Stachybotrys atra by substrate analogues,” Carbohydrate Research, vol. 138, pp. 127–134, 1985.
- V. Gopalan, L. B. Daniels, R. H. Glew, and M. Claeyssens, “Kinetic analysis of the interaction of alkyl glycosides with two human β-glucosidases,” Biochemical Journal, vol. 262, no. 2, pp. 541–548, 1989.
- M. S. Cheng, Q. L. Wang, Q. Tian et al., “Total synthesis of methyl protodioscin: a potent agent with antitumor activity,” The Journal of Organic Chemistry, vol. 68, no. 9, pp. 3658–3662, 2003.
- S. Weng, “Diastereoselective thioglycosylation of peracetylated glycosides catalyzed by in situ generated iron(III) iodide from elemental iodine and iron,” Tetrahedron Letters, vol. 50, no. 46, pp. 6414–6417, 2009.
- J. Kuhn, E. M. Pettersson, B. K. Feld et al., “Sequestration of plant-derived phenolglucosides by larvae of the leaf beetle Chrysomela lapponica: thioglucosides as mechanistic probes,” Journal of Chemical Ecology, vol. 33, no. 1, pp. 5–24, 2007.
- E. M. Montgomery, N. K. Richtmyer, and C. S. Hudson, “Attempts to find new antimalarials. VIII. Phenyl β-D-glucothiosides, diphenyl disulfides, phenyl thiocyanates, and related compounds,” Journal of Organic Chemistry, vol. 11, no. 3, pp. 301–306, 1946.
- W. T. Haskins, R. M. Hann, and C. S. Hudson, “Relations between rotatory power and structure in the sugar group. XXXV. Some 2′-naphthyl 1-thioglycopyranosides and their acetates,” Journal of the American Chemical Society, vol. 69, no. 7, pp. 1668–1672, 1947.
- S. A. Galema, J. B. F. N. Engberts, and H. A. van Doren, “Synthesis, purification and liquid-crystalline behaviour of several alkyl 1-thio-D-glycopyranosides,” Carbohydrate Research, vol. 303, no. 4, pp. 423–434, 1997.
- H. Lineweaver and D. Burk, “The determination of enzyme dissociation constants,” Journal of the American Chemical Society, vol. 56, no. 3, pp. 658–666, 1934.
- I. H. Segel, Biochemical Calculations, John Wiley & Sons, New York, NY, USA, 1976.
Copyright © 2014 Andrey V. Samoshin et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.